Addgene TRRAP sgRNA3 - Sequence Analyzer Skip to main content
Addgene

Sequence Analyzer: TRRAP sgRNA3


Map View

SnaBI (12,988) SpeI (12,647) pRS-marker (12,442 .. 12,461) SgrDI (12,396) AmpR promoter SspI (12,279) Amp-R (12,025 .. 12,044) PvuI (11,845) FspI (11,697) pBR322ori-F (10,723 .. 10,742) PciI (10,582) L4440 (10,472 .. 10,489) CAP binding site lac promoter M13/pUC Reverse (10,254 .. 10,276) lac operator M13 Reverse (10,241 .. 10,257) M13 rev BstZ17I (10,203) EBV-rev (10,162 .. 10,181) SV40pA-R (10,108 .. 10,127) FseI (9843) SgrAI (9681) MauBI (9603) EM7 promoter AvrII (9438) SV40pro-F (9366 .. 9385) pBABE 3' (9119 .. 9139) F1ori-F (8979 .. 9000) F1ori-R (8769 .. 8788) BGH-rev (8406 .. 8423) PmeI (8390) WPRE-R (7540 .. 7560) Puro-F (7368 .. 7388) BstEII (7006) RsrII (6988) BsiWI (6928) PflFI - Tth111I (6914) BamHI (6809) CMV promoter CMV-F (154 .. 174) 5' LTR (truncated) NotI (907) BbvCI (1183) AleI (1336) gp41 peptide cPPT/CTS PacI (1980) Acc65I (1984) KpnI (1988) hU6-F (1990 .. 2010) LKO.1 5' (2161 .. 2180) EcoRI (2342) NheI (2348) BmtI (2352) AgeI (2610) XbaI (2616) AfeI (2623) EcoRV (4486) BsaBI * (6131) FLAG TRRAP sgRNA3 13,013 bp

Sequence View

5ʹ 3ʹ 60 CMV promoter t g a t g c g g t t t t g g c a g t a c a t c a a t g g g c g t g g a t a g c g g t t t g a c t c a c g g g g a t t t c a c t a c g c c a a a a c c g t c a t g t a g t t a c c c g c a c c t a t c g c c a a a c t g a g t g c c c c t a a a g

120 CMV promoter c a a g t c t c c a c c c c a t t g a c g t c a a t g g g a g t t t g t t t t g g c a c c a a a a t c a a c g g g a c t g t t c a g a g g t g g g g t a a c t g c a g t t a c c c t c a a a c a a a a c c g t g g t t t t a g t t g c c c t g a

180 CMV promoter C G C A A A T G G G C G G T A G G C G T G CMV-F t t c c a a a a t g t c g t a a c a a c t c c g c c c c a t t g a c g c a a a t g g g c g g t a g g c g t g t a c g g t a a g g t t t t a c a g c a t t g t t g a g g c g g g g t a a c t g c g t t t a c c c g c c a t c c g c a c a t g c c a

240 CMV promoter 5' LTR (truncated) g g g a g g t c t a t a t a a g c a g c g c g t t t t g c c t g t a c t g g g t c t c t c t g g t t a g a c c a g a t c c c c t c c a g a t a t a t t c g t c g c g c a a a a c g g a c a t g a c c c a g a g a g a c c a a t c t g g t c t a g

300 5' LTR (truncated) t g a g c c t g g g a g c t c t c t g g c t a a c t a g g g a a c c c a c t g c t t a a g c c t c a a t a a a g c t t g a c t c g g a c c c t c g a g a g a c c g a t t g a t c c c t t g g g t g a c g a a t t c g g a g t t a t t t c g a a c

360 5' LTR (truncated) c c t t g a g t g c t t c a a g t a g t g t g t g c c c g t c t g t t g t g t g a c t c t g g t a a c t a g a g a t c c g g a a c t c a c g a a g t t c a t c a c a c a c g g g c a g a c a a c a c a c t g a g a c c a t t g a t c t c t a g g

420 5' LTR (truncated) c t c a g a c c c t t t t a g t c a g t g t g g a a a a t c t c t a g c a g t g g c g c c c g a a c a g g g a c t t g a g a g t c t g g g a a a a t c a g t c a c a c c t t t t a g a g a t c g t c a c c g c g g g c t t g t c c c t g a a c t

480 HIV-1 Ψ a a g c g a a a g g g a a a c c a g a g g a g c t c t c t c g a c g c a g g a c t c g g c t t g c t g a a g c g c g c a t t c g c t t t c c c t t t g g t c t c c t c g a g a g a g c t g c g t c c t g a g c c g a a c g a c t t c g c g c g t

540 HIV-1 Ψ c g g c a a g a g g c g a g g g g c g g c g a c t g g t g a g t a c g c c a a a a a t t t t g a c t a g c g g a g g c t g c c g t t c t c c g c t c c c c g c c g c t g a c c a c t c a t g c g g t t t t t a a a a c t g a t c g c c t c c g a

600 HIV-1 Ψ a g a a g g a g a g a g a t g g g t g c g a g a g c g t c a g t a t t a a g c g g g g g a g a a t t a g a t c g c g a t t c t t c c t c t c t c t a c c c a c g c t c t c g c a g t c a t a a t t c g c c c c c t c t t a a t c t a g c g c t a

660 g g g a a a a a a t t c g g t t a a g g c c a g g g g g a a a g a a a a a a t a t a a a t t a a a a c a t a t a g t a t c c c t t t t t t a a g c c a a t t c c g g t c c c c c t t t c t t t t t t a t a t t t a a t t t t g t a t a t c a t a

720 g g g c a a g c a g g g a g c t a g a a c g a t t c g c a g t t a a t c c t g g c c t g t t a g a a a c a t c a g a a g c c c g t t c g t c c c t c g a t c t t g c t a a g c g t c a a t t a g g a c c g g a c a a t c t t t g t a g t c t t c

780 g c t g t a g a c a a a t a c t g g g a c a g c t a c a a c c a t c c c t t c a g a c a g g a t c a g a a g a a c t t a c g a c a t c t g t t t a t g a c c c t g t c g a t g t t g g t a g g g a a g t c t g t c c t a g t c t t c t t g a a t

840 g a t c a t t a t a t a a t a c a g t a g c a a c c c t c t a t t g t g t g c a t c a a a g g a t a g a g a t a a a a g c t a g t a a t a t a t t a t g t c a t c g t t g g g a g a t a a c a c a c g t a g t t t c c t a t c t c t a t t t t c

900 a c a c c a a g g a a g c t t t a g a c a a g a t a g a g g a a g a g c a a a a c a a a a g t a a g a c c a c c g c a c t g t g g t t c c t t c g a a a t c t g t t c t a t c t c c t t c t c g t t t t g t t t t c a t t c t g g t g g c g t g

NotI 960 MRDNWRS a g c a a g c g g c c g c t g a t c t t c a g a c c t g g a g g a g g a g a t a t g a g g g a c a a t t g g a g a a g t t c g t t c g c c g g c g a c t a g a a g t c t g g a c c t c c t c c t c t a t a c t c c c t g t t a a c c t c t t c a

1020 ELYKYKVVKIEPLGVAPTKA g a a t t a t a t a a a t a t a a a g t a g t a a a a a t t g a a c c a t t a g g a g t a g c a c c c a c c a a g g c a c t t a a t a t a t t t a t a t t t c a t c a t t t t t a a c t t g g t a a t c c t c a t c g t g g g t g g t t c c g t

1080 RRE KRRVVQREKRAVGIGALFLG a a g a g a a g a g t g g t g c a g a g a g a a a a a a g a g c a g t g g g a a t a g g a g c t t t g t t c c t t g g g t t c t c t t c t c a c c a c g t c t c t c t t t t t t c t c g t c a c c c t t a t c c t c g a a a c a a g g a a c c c

1140 RRE FLGAAGSTMGAASMTLTVQA t t c t t g g g a g c a g c a g g a a g c a c t a t g g g c g c a g c g t c a a t g a c g c t g a c g g t a c a g g c c a a g a a c c c t c g t c g t c c t t c g t g a t a c c c g c g t c g c a g t t a c t g c g a c t g c c a t g t c c g g

BbvCI 1200 RRE RQLLSGIVQQQNNLLRAIEA a g a c a a t t a t t g t c t g g t a t a g t g c a g c a g c a g a a c a a t t t g c t g a g g g c t a t t g a g g c g t c t g t t a a t a a c a g a c c a t a t c a c g t c g t c g t c t t g t t a a a c g a c t c c c g a t a a c t c c g c

1260 RRE QQHLLQLTVWGIKQLQARIL c a a c a g c a t c t g t t g c a a c t c a c a g t c t g g g g c a t c a a g c a g c t c c a g g c a a g a a t c c t g g t t g t c g t a g a c a a c g t t g a g t g t c a g a c c c c g t a g t t c g t c g a g g t c c g t t c t t a g g a c

1320 RRE AVERYLKDQQLLGIWGCSGK g c t g t g g a a a g a t a c c t a a a g g a t c a a c a g c t c c t g g g g a t t t g g g g t t g c t c t g g a a a a c g a c a c c t t t c t a t g g a t t t c c t a g t t g t c g a g g a c c c c t a a a c c c c a a c g a g a c c t t t t

AleI 1380 LICTTAVPWNASWSNKSLEQ c t c a t t t g c a c c a c t g c t g t g c c t t g g a a t g c t a g t t g g a g t a a t a a a t c t c t g g a a c a g g a g t a a a c g t g g t g a c g a c a c g g a a c c t t a c g a t c a a c c t c a t t a t t t a g a g a c c t t g t c

1440 IWNHTTWMEWDREINNYTSL a t t t g g a a t c a c a c g a c c t g g a t g g a g t g g g a c a g a g a a a t t a a c a a t t a c a c a a g c t t a t a a a c c t t a g t g t g c t g g a c c t a c c t c a c c c t g t c t c t t t a a t t g t t a a t g t g t t c g a a t

1500 1 5 KNEQELL gp41 peptide IHSLIEESQNQQEKNEQELL a t a c a c t c c t t a a t t g a a g a a t c g c a a a a c c a g c a a g a a a a g a a t g a a c a a g a a t t a t t g t a t g t g a g g a a t t a a c t t c t t a g c g t t t t g g t c g t t c t t t t c t t a c t t g t t c t t a a t a a c

1560 10 15 ELDKWASL (in frame with gp41 peptide) WNWFNITNWLWY gp41 peptide ELDKWASLWNWFNITNWLWY g a a t t a g a t a a a t g g g c a a g t t t g t g g a a t t g g t t t a a c a t a a c a a a t t g g c t g t g g t a t c t t a a t c t a t t t a c c c g t t c a a a c a c c t t a a c c a a a t t g t a t t g t t t a a c c g a c a c c a t a

1620 (in frame with gp41 peptide) IKLFIMIVGGLVGLRIVFAV IKLFIMIVGGLVGLRIVFAV a t a a a a t t a t t c a t a a t g a t a g t a g g a g g c t t g g t a g g t t t a a g a a t a g t t t t t g c t g t a t a t t t t a a t a a g t a t t a c t a t c a t c c t c c g a a c c a t c c a a a t t c t t a t c a a a a a c g a c a t

1680 (in frame with gp41 peptide) LSIVNRVRQGYSPLSFQTHL LSIVNRVRQGYSPLSFQTHL c t t t c t a t a g t g a a t a g a g t t a g g c a g g g a t a t t c a c c a t t a t c g t t t c a g a c c c a c c t c g a a a g a t a t c a c t t a t c t c a a t c c g t c c c t a t a a g t g g t a a t a g c a a a g t c t g g g t g g a g

1740 (in frame with gp41 peptide) PTPRGPDRPEGIEEEGGERD PTPRGPDRPEGIEEEGGERD c c a a c c c c g a g g g g a c c c g a c a g g c c c g a a g g a a t a g a a g a a g a a g g t g g a g a g a g a g a c g g t t g g g g c t c c c c t g g g c t g t c c g g g c t t c c t t a t c t t c t t c t t c c a c c t c t c t c t c t g

1800 (in frame with gp41 peptide) RDRSIRLVNGSALRAPILQT RDRSIRLVNGSALRAPILQT a g a g a c a g a t c c a t t c g a t t a g t g a a c g g a t c g g c a c t g c g t g c g c c a a t t c t g c a g a c a t c t c t g t c t a g g t a a g c t a a t c a c t t g c c t a g c c g t g a c g c a c g c g g t t a a g a c g t c t g t

1860 (in frame with gp41 peptide) NGSIHPQF* cPPT/CTS NGSIHPQF* a a t g g c a g t a t t c a t c c a c a a t t t t a a a a g a a a a g g g g g g a t t g g g g g g t a c a g t g c a g g t t a c c g t c a t a a g t a g g t g t t a a a a t t t t c t t t t c c c c c c t a a c c c c c c a t g t c a c g t c c

1920 cPPT/CTS g g a a a g a a t a g t a g a c a t a a t a g c a a c a g a c a t a c a a a c t a a a g a a t t a c a a a a a c a a a t c c t t t c t t a t c a t c t g t a t t a t c g t t g t c t g t a t g t t t g a t t t c t t a a t g t t t t t g t t t a

PacI 1980 cPPT/CTS t a c a a a a a t t c a a a a t t t t c g g g t t t a t t a c a g g g a c a g c a g a g a t c c a g t t t g g t t a a t a t g t t t t t a a g t t t t a a a a g c c c a a a t a a t g t c c c t g t c g t c t c t a g g t c a a a c c a a t t a

Acc65IKpnI 2040 U6 promoter G A G G G C C T A T T T C C C A T G A T T hU6-F t a a g g t a c c g a g g g c c t a t t t c c c a t g a t t c c t t c a t a t t t g c a t a t a c g a t a c a a g g c t a t t c c a t g g c t c c c g g a t a a a g g g t a c t a a g g a a g t a t a a a c g t a t a t g c t a t g t t c c g a

2100 U6 promoter g t t a g a g a g a t a a t t a g a a t t a a t t t g a c t g t a a a c a c a a a g a t a t t a g t a c a a a a t a c g c a a t c t c t c t a t t a a t c t t a a t t a a a c t g a c a t t t g t g t t t c t a t a a t c a t g t t t t a t g c

2160 U6 promoter t g a c g t a g a a a g t a a t a a t t t c t t g g g t a g t t t g c a g t t t t a a a a t t a t g t t t t a a a a t g a c t g c a t c t t t c a t t a t t a a a g a a c c c a t c a a a c g t c a a a a t t t t a a t a c a a a a t t t t a c

2220 U6 promoter G A C T A T C A T A T G C T T A C C G T LKO.1 5' g a c t a t c a t a t g c t t a c c g t a a c t t g a a a g t a t t t c g a t t t c t t g g c t t t a t a t a t c t t g c t g a t a g t a t a c g a a t g g c a t t g a a c t t t c a t a a a g c t a a a g a a c c g a a a t a t a t a g a a c

2280 U6 promoter gRNA scaffold t g g a a a g g a c g a a a c a c c g c c a c t g g g g a t c g t t c a g t g g t t t t a g a g c t a g a a a t a g c a a c c t t t c c t g c t t t g t g g c g g t g a c c c c t a g c a a g t c a c c a a a a t c t c g a t c t t t a t c g t

2340 gRNA scaffold *DTILKFFHCRTPAKK a g t t a a a a t a a g g c t a g t c c g t t a t c a a c t t g a a a a a g t g g c a c c g a g t c g g t g c t t t t t t c a a t t t t a t t c c g a t c a g g c a a t a g t t g a a c t t t t t c a c c g t g g c t c a g c c a c g a a a a a

BmtINheIEcoRI 2400 EF-1 α core promoter FESALDQFSHSNAGTGTLPC t g a a t t c g c t a g c t a g g t c t t g a a a g g a g t g g g a a t t g g c t c c g g t g c c c g t c a g t g g g c a c t t a a g c g a t c g a t c c a g a a c t t t c c t c a c c c t t a a c c g a g g c c a c g g g c a g t c a c c c g

2460 EF-1 α core promoter LACRGCDGLLQPSPDAISGT a g a g c g c a c a t c g c c c a c a g t c c c c g a g a a g t t g g g g g g a g g g g t c g g c a a t t g a t c c g g t c t c g c g t g t a g c g g g t g t c a g g g g c t c t t c a a c c c c c c t c c c c a g c c g t t a a c t a g g c c

2520 EF-1 α core promoter GLSPPAPYVPFTIDHVPEAK t g c c t a g a g a a g g t g g c g c g g g g t a a a c t g g g a a a g t g a t g t c g t g t a c t g g c t c c g c c t a c g g a t c t c t t c c a c c g c g c c c c a t t t g a c c c t t t c a c t a c a g c a c a t g a c c g a g g c g g a

2580 EF-1 α core promoter KGLTPSFRIYTCYDGHVNKK t t t t c c c g a g g g t g g g g g a g a a c c g t a t a t a a g t g c a g t a g t c g c c g t g a a c g t t c t t t t a a a a g g g c t c c c a c c c c c t c t t g g c a t a t a t t c a c g t c a t c a g c g g c a c t t g c a a g a a a a

AfeIXbaIAgeI 2640 1 MDK EF-1 α core promoter Cas9 1 MDK Cas9 Kozak sequence MDK AVPKGGSCLVPELASGGHVL t c g c a a c g g g t t t g c c g c c a g a a c a c a g g a c c g g t t c t a g a g c g c t g c c a c c a t g g a c a a a g c g t t g c c c a a a c g g c g g t c t t g t g t c c t g g c c a a g a t c t c g c g a c g g t g g t a c c t g t t

2700 5 10 15 20 KYSIGLDIGTNSVGWAVITD Cas9 5 10 15 20 KYSIGLDIGTNSVGWAVITD Cas9 KYSIGLDIGTNSVGWAVITD LVADAQVDAGVRHAPGHDGV g a a g t a c a g c a t c g g c c t g g a c a t c g g c a c c a a c t c t g t g g g c t g g g c c g t g a t c a c c g a c t t c a t g t c g t a g c c g g a c c t g t a g c c g t g g t t g a g a c a c c c g a c c c g g c a c t a g t g g c t

2760 25 30 35 40 EYKVPSKKFKVLGNTDRHSI Cas9 25 30 35 40 EYKVPSKKFKVLGNTDRHSI Cas9 EYKVPSKKFKVLGNTDRHSI LVLHGALFELHQAVGVPVAD c g a g t a c a a g g t g c c c a g c a a g a a a t t c a a g g t g c t g g g c a a c a c c g a c c g g c a c a g c a t g c t c a t g t t c c a c g g g t c g t t c t t t a a g t t c c a c g a c c c g t t g t g g c t g g c c g t g t c g t a

2820 45 50 55 60 KKNLIGALLFDSGETAEATR Cas9 45 50 55 60 KKNLIGALLFDSGETAEATR Cas9 KKNLIGALLFDSGETAEATR LLVQDSGQQEVAAFCGLGGP c a a g a a g a a c c t g a t c g g a g c c c t g c t g t t c g a c a g c g g c g a a a c a g c c g a g g c c a c c c g g t t c t t c t t g g a c t a g c c t c g g g a c g a c a a g c t g t c g c c g c t t t g t c g g c t c c g g t g g g c

2880 65 70 75 80 LKRTARRRYTRRKNRICYLQ Cas9 65 70 75 80 LKRTARRRYTRRKNRICYLQ Cas9 LKRTARRRYTRRKNRICYLQ QLSGGSSSVGSPLVPDAIQL g c t g a a g a g a a c c g c c a g a a g a a g a t a c a c c a g a c g g a a g a a c c g g a t c t g c t a t c t g c a c g a c t t c t c t t g g c g g t c t t c t t c t a t g t g g t c t g c c t t c t t g g c c t a g a c g a t a g a c g t

2940 85 90 95 100 EIFSNEMAKVDDSFFHRLEE Cas9 85 90 95 100 EIFSNEMAKVDDSFFHRLEE Cas9 EIFSNEMAKVDDSFFHRLEE LDEAVLHGLHVVAEEVSQFL a g a g a t c t t c a g c a a c g a g a t g g c c a a g g t g g a c g a c a g c t t c t t c c a c a g a c t g g a a g a t c t c t a g a a g t c g t t g c t c t a c c g g t t c c a c c t g c t g t c g a a g a a g g t g t c t g a c c t t c t

3000 105 110 115 120 SFLVEEDKKHERHPIFGNIV Cas9 105 110 115 120 SFLVEEDKKHERHPIFGNIV Cas9 SFLVEEDKKHERHPIFGNIV GEQHFLILLVLPVGDEAVDH g t c c t t c c t g g t g g a a g a g g a t a a g a a g c a c g a g c g g c a c c c c a t c t t c g g c a a c a t c g t c a g g a a g g a c c a c c t t c t c c t a t t c t t c g t g c t c g c c g t g g g g t a g a a g c c g t t g t a g c a

3060 125 130 135 140 DEVAYHEKYPTIYHLRKKLV Cas9 125 130 135 140 DEVAYHEKYPTIYHLRKKLV Cas9 DEVAYHEKYPTIYHLRKKLV VLHGVVLLVGGDVVQSLFQH g g a c g a g g t g g c c t a c c a c g a g a a g t a c c c c a c c a t c t a c c a c c t g a g a a a g a a a c t g g t c c t g c t c c a c c g g a t g g t g c t c t t c a t g g g g t g g t a g a t g g t g g a c t c t t t c t t t g a c c a

3120 145 150 155 160 DSTDKADLRLIYLALAHMIK Cas9 145 150 155 160 DSTDKADLRLIYLALAHMIK Cas9 DSTDKADLRLIYLALAHMIK VAGVLGVQPQDIQGQGVHDL g g a c a g c a c c g a c a a g g c c g a c c t g c g g c t g a t c t a t c t g g c c c t g g c c c a c a t g a t c a a c c t g t c g t g g c t g t t c c g g c t g g a c g c c g a c t a g a t a g a c c g g g a c c g g g t g t a c t a g t t

3180 165 170 175 180 FRGHFLIEGDLNPDNSDVDK Cas9 165 170 175 180 FRGHFLIEGDLNPDNSDVDK Cas9 FRGHFLIEGDLNPDNSDVDK EPAVEQDLAVQVGVVAVHVL g t t c c g g g g c c a c t t c c t g a t c g a g g g c g a c c t g a a c c c c g a c a a c a g c g a c g t g g a c a a c a a g g c c c c g g t g a a g g a c t a g c t c c c g c t g g a c t t g g g g c t g t t g t c g c t g c a c c t g t t

3240 185 190 195 200 LFIQLVQTYNQLFEENPINA Cas9 185 190 195 200 LFIQLVQTYNQLFEENPINA Cas9 LFIQLVQTYNQLFEENPINA QEDLQHLGVVLQELFVGDVG g c t g t t c a t c c a g c t g g t g c a g a c c t a c a a c c a g c t g t t c g a g g a a a a c c c c a t c a a c g c c g a c a a g t a g g t c g a c c a c g t c t g g a t g t t g g t c g a c a a g c t c c t t t t g g g g t a g t t g c g

3300 205 210 215 220 SGVDAKAILSARLSKSRRLE Cas9 205 210 215 220 SGVDAKAILSARLSKSRRLE Cas9 SGVDAKAILSARLSKSRRLE AAHVGLGDQRGSQALASPQF c a g c g g c g t g g a c g c c a a g g c c a t c c t g t c t g c c a g a c t g a g c a a g a g c a g a c g g c t g g a g t c g c c g c a c c t g c g g t t c c g g t a g g a c a g a c g g t c t g a c t c g t t c t c g t c t g c c g a c c t

3360 225 230 235 240 NLIAQLPGEKKNGLFGNLIA Cas9 225 230 235 240 NLIAQLPGEKKNGLFGNLIA Cas9 NLIAQLPGEKKNGLFGNLIA IQDGLQGALLLIAQESVQNG a a a t c t g a t c g c c c a g c t g c c c g g c g a g a a g a a g a a t g g c c t g t t c g g a a a c c t g a t t g c t t t a g a c t a g c g g g t c g a c g g g c c g c t c t t c t t c t t a c c g g a c a a g c c t t t g g a c t a a c g

3420 245 250 255 260 LSLGLTPNFKSNFDLAEDAK Cas9 245 250 255 260 LSLGLTPNFKSNFDLAEDAK Cas9 LSLGLTPNFKSNFDLAEDAK QAQAQGGVELAVEVQGLIGF c c t g a g c c t g g g c c t g a c c c c c a a c t t c a a g a g c a a c t t c g a c c t g g c c g a g g a t g c c a a g g a c t c g g a c c c g g a c t g g g g g t t g a a g t t c t c g t t g a a g c t g g a c c g g c t c c t a c g g t t

3480 265 270 275 280 LQLSKDTYDDDLDNLLAQIG Cas9 265 270 275 280 LQLSKDTYDDDLDNLLAQIG Cas9 LQLSKDTYDDDLDNLLAQIG QLQALVGVVVVQVVQQGLDA a c t g c a g c t g a g c a a g g a c a c c t a c g a c g a c g a c c t g g a c a a c c t g c t g g c c c a g a t c g g t g a c g t c g a c t c g t t c c t g t g g a t g c t g c t g c t g g a c c t g t t g g a c g a c c g g g t c t a g c c

3540 285 290 295 300 DQYADLFLAAKNLSDAILLS Cas9 285 290 295 300 DQYADLFLAAKNLSDAILLS Cas9 DQYADLFLAAKNLSDAILLS VLVGVQKQGGLVQGVGDQQA c g a c c a g t a c g c c g a c c t g t t t c t g g c c g c c a a g a a c c t g t c c g a c g c c a t c c t g c t g a g g c t g g t c a t g c g g c t g g a c a a a g a c c g g c g g t t c t t g g a c a g g c t g c g g t a g g a c g a c t c

3600 305 310 315 320 DILRVNTEITKAPLSASMIK Cas9 305 310 315 320 DILRVNTEITKAPLSASMIK Cas9 DILRVNTEITKAPLSASMIK VDQSHVGLDGLGGQAGRHDL c g a c a t c c t g a g a g t g a a c a c c g a g a t c a c c a a g g c c c c c c t g a g c g c c t c t a t g a t c a a g c t g t a g g a c t c t c a c t t g t g g c t c t a g t g g t t c c g g g g g g a c t c g c g g a g a t a c t a g t t

3660 325 330 335 340 RYDEHHQDLTLLKALVRQQL Cas9 325 330 335 340 RYDEHHQDLTLLKALVRQQL Cas9 RYDEHHQDLTLLKALVRQQL SVVLVVLVQGQQFSEHPLLQ g a g a t a c g a c g a g c a c c a c c a g g a c c t g a c c c t g c t g a a a g c t c t c g t g c g g c a g c a g c t c t c t a t g c t g c t c g t g g t g g t c c t g g a c t g g g a c g a c t t t c g a g a g c a c g c c g t c g t c g a

3720 345 350 355 360 PEKYKEIFFDQSKNGYAGYI Cas9 345 350 355 360 PEKYKEIFFDQSKNGYAGYI Cas9 PEKYKEIFFDQSKNGYAGYI RLLVFLNEEVLALVAVGAVN g c c t g a g a a g t a c a a a g a g a t t t t c t t c g a c c a g a g c a a g a a c g g c t a c g c c g g c t a c a t c g g a c t c t t c a t g t t t c t c t a a a a g a a g c t g g t c t c g t t c t t g c c g a t g c g g c c g a t g t a

3780 365 370 375 380 DGGASQEEFYKFIKPILEKM Cas9 365 370 375 380 DGGASQEEFYKFIKPILEKM Cas9 DGGASQEEFYKFIKPILEKM VASGALFLEVLEDLGDQFLH t g a c g g c g g a g c c a g c c a g g a a g a g t t c t a c a a g t t c a t c a a g c c c a t c c t g g a a a a g a t a c t g c c g c c t c g g t c g g t c c t t c t c a a g a t g t t c a a g t a g t t c g g g t a g g a c c t t t t c t a

3840 385 390 395 400 DGTEELLVKLNREDLLRKQR Cas9 385 390 395 400 DGTEELLVKLNREDLLRKQR Cas9 DGTEELLVKLNREDLLRKQR VAGLFQEHLQVSLVQQPLLP g g a c g g c a c c g a g g a a c t g c t c g t g a a g c t g a a c a g a g a g g a c c t g c t g c g g a a g c a g c g c c t g c c g t g g c t c c t t g a c g a g c a c t t c g a c t t g t c t c t c c t g g a c g a c g c c t t c g t c g c

3900 405 410 415 420 TFDNGSIPHQIHLGELHAIL Cas9 405 410 415 420 TFDNGSIPHQIHLGELHAIL Cas9 TFDNGSIPHQIHLGELHAIL GEVVAADGVLDVQSLQVGNQ g a c c t t c g a c a a c g g c a g c a t c c c c c a c c a g a t c c a c c t g g g a g a g c t g c a c g c c a t t c t c t g g a a g c t g t t g c c g t c g t a g g g g g t g g t c t a g g t g g a c c c t c t c g a c g t g c g g t a a g a

3960 425 430 435 440 RRQEDFYPFLKDNREKIEKI Cas9 425 430 435 440 RRQEDFYPFLKDNREKIEKI Cas9 RRQEDFYPFLKDNREKIEKI PPLFIKVWEQLVVPFLDLLD g c g g c g g c a g g a a g a t t t t t a c c c a t t c c t g a a g g a c a a c c g g g a a a a g a t c g a g a a g a t c g c c g c c g t c c t t c t a a a a a t g g g t a a g g a c t t c c t g t t g g c c c t t t t c t a g c t c t t c t a

4020 445 450 455 460 LTFRIPYYVGPLARGNSRFA Cas9 445 450 455 460 LTFRIPYYVGPLARGNSRFA Cas9 LTFRIPYYVGPLARGNSRFA QGEADGVVHARQGPSVASEG c c t g a c c t t c c g c a t c c c c t a c t a c g t g g g c c c t c t g g c c a g g g g a a a c a g c a g a t t c g c g g a c t g g a a g g c g t a g g g g a t g a t g c a c c c g g g a g a c c g g t c c c c t t t g t c g t c t a a g c g

4080 465 470 475 480 WMTRKSEETITPWNFEEVVD Cas9 465 470 475 480 WMTRKSEETITPWNFEEVVD Cas9 WMTRKSEETITPWNFEEVVD PHGSLALFGDGGPVELFHHV c t g g a t g a c c a g a a a g a g c g a g g a a a c c a t c a c c c c c t g g a a c t t c g a g g a a g t g g t g g a g a c c t a c t g g t c t t t c t c g c t c c t t t g g t a g t g g g g g a c c t t g a a g c t c c t t c a c c a c c t

4140 485 490 495 500 KGASAQSFIERMTNFDKNLP Cas9 485 490 495 500 KGASAQSFIERMTNFDKNLP Cas9 KGASAQSFIERMTNFDKNLP LASGGLAEDLPHGVEILVQG c a a g g g c g c t t c c g c c c a g a g c t t c a t c g a g c g g a t g a c c a a c t t c g a t a a g a a c c t g c c g t t c c c g c g a a g g c g g g t c t c g a a g t a g c t c g c c t a c t g g t t g a a g c t a t t c t t g g a c g g

4200 505 510 515 520 NEKVLPKHSLLYEYFTVYNE Cas9 505 510 515 520 NEKVLPKHSLLYEYFTVYNE Cas9 NEKVLPKHSLLYEYFTVYNE VLLHQGLVAQQVLVEGHIVL c a a c g a g a a g g t g c t g c c c a a g c a c a g c c t g c t g t a c g a g t a c t t c a c c g t g t a t a a c g a g t t g c t c t t c c a c g a c g g g t t c g t g t c g g a c g a c a t g c t c a t g a a g t g g c a c a t a t t g c t

4260 525 530 535 540 LTKVKYVTEGMRKPAFLSGE Cas9 525 530 535 540 LTKVKYVTEGMRKPAFLSGE Cas9 LTKVKYVTEGMRKPAFLSGE QGFHFVHGLSHSLGGEQAAL g c t g a c c a a a g t g a a a t a c g t g a c c g a g g g a a t g a g a a a g c c c g c c t t c c t g a g c g g c g a c g a c t g g t t t c a c t t t a t g c a c t g g c t c c c t t a c t c t t t c g g g c g g a a g g a c t c g c c g c t

4320 545 550 555 560 QKKAIVDLLFKTNRKVTVKQ Cas9 545 550 555 560 QKKAIVDLLFKTNRKVTVKQ Cas9 QKKAIVDLLFKTNRKVTVKQ LFLGDHVQQELGVPFHGHLL g c a g a a a a a g g c c a t c g t g g a c c t g c t g t t c a a g a c c a a c c g g a a a g t g a c c g t g a a g c a c g t c t t t t t c c g g t a g c a c c t g g a c g a c a a g t t c t g g t t g g c c t t t c a c t g g c a c t t c g t

4380 565 570 575 580 LKEDYFKKIECFDSVEISGV Cas9 565 570 575 580 LKEDYFKKIECFDSVEISGV Cas9 LKEDYFKKIECFDSVEISGV QFLVVELFDLAEVGHFDGAH g c t g a a a g a g g a c t a c t t c a a g a a a a t c g a g t g c t t c g a c t c c g t g g a a a t c t c c g g c g t c g a c t t t c t c c t g a t g a a g t t c t t t t a g c t c a c g a a g c t g a g g c a c c t t t a g a g g c c g c a

4440 585 590 595 600 EDRFNASLGTYHDLLKIIKD Cas9 585 590 595 600 EDRFNASLGTYHDLLKIIKD Cas9 EDRFNASLGTYHDLLKIIKD FIPEVGGQACVVIQQFNDLV g g a a g a t c g g t t c a a c g c c t c c c t g g g c a c a t a c c a c g a t c t g c t g a a a a t t a t c a a g g a c c t t c t a g c c a a g t t g c g g a g g g a c c c g t g t a t g g t g c t a g a c g a c t t t t a a t a g t t c c t

EcoRV 4500 605 610 615 620 KDFLDNEENEDILEDIVLTL Cas9 605 610 615 620 KDFLDNEENEDILEDIVLTL Cas9 KDFLDNEENEDILEDIVLTL LVEQVILFVLVNQFIDHQGQ c a a g g a c t t c c t g g a c a a t g a g g a a a a c g a g g a c a t t c t g g a a g a t a t c g t g c t g a c c c t g t t c c t g a a g g a c c t g t t a c t c c t t t t g c t c c t g t a a g a c c t t c t a t a g c a c g a c t g g g a

4560 625 630 635 640 TLFEDREMIEERLKTYAHLF Cas9 625 630 635 640 TLFEDREMIEERLKTYAHLF Cas9 TLFEDREMIEERLKTYAHLF *AWRN CQKLVSLHDLFPQFGIGVQE g a c a c t g t t t g a g g a c a g a g a g a t g a t c g a g g a a c g g c t g a a a a c c t a t g c c c a c c t g t t c t g t g a c a a a c t c c t g t c t c t c t a c t a g c t c c t t g c c g a c t t t t g g a t a c g g g t g g a c a a

4620 645 650 655 660 DDKVMKQLKRRRYTGWGRLS Cas9 645 650 655 660 DDKVMKQLKRRRYTGWGRLS Cas9 DDKVMKQLKRRRYTGWGRLS SSLTIFCSFRRLYVPQPLSL VVFHHLLQLPPSVGAPAPQA c g a c g a c a a a g t g a t g a a g c a g c t g a a g c g g c g g a g a t a c a c c g g c t g g g g c a g g c t g a g g c t g c t g t t t c a c t a c t t c g t c g a c t t c g c c g c c t c t a t g t g g c c g a c c c c g t c c g a c t c

4680 665 670 675 680 RKLINGIRDKQSGKTILDFL Cas9 665 670 675 680 RKLINGIRDKQSGKTILDFL Cas9 RKLINGIRDKQSGKTILDFL RFSILPMRSLCDPLVIRSKR PLQDVADPVLLGALCDQIEQ c c g g a a g c t g a t c a a c g g c a t c c g g g a c a a g c a g t c c g g c a a g a c a a t c c t g g a t t t c c t g g c c t t c g a c t a g t t g c c g t a g g c c c t g t t c g t c a g g c c g t t c t g t t a g g a c c t a a a g g a

4740 685 690 695 700 KSDGFANRNFMQLIHDDSLT Cas9 685 690 695 700 KSDGFANRNFMQLIHDDSLT Cas9 KSDGFANRNFMQLIHDDSLT FDSPKALLFKMCSIWSSLRV LGVAEGVSVEHLQDVVVAQG g a a g t c c g a c g g c t t c g c c a a c a g a a a c t t c a t g c a g c t g a t c c a c g a c g a c a g c c t g a c c t t c a g g c t g c c g a a g c g g t t g t c t t t g a a g t a c g t c g a c t a g g t g c t g c t g t c g g a c t g

4800 705 710 715 720 FKEDIQKAQVSGQGDSLHEH Cas9 705 710 715 720 FKEDIQKAQVSGQGDSLHEH Cas9 FKEDIQKAQVSGQGDSLHEH KLSSMWFAWTDPWPSLRCSC KFLVDLFGLHGALAIAQVLV c t t t a a a g a g g a c a t c c a g a a a g c c c a g g t g t c c g g c c a g g g c g a t a g c c t g c a c g a g c a g a a a t t t c t c c t g t a g g t c t t t c g g g t c c a c a g g c c g g t c c c g c t a t c g g a c g t g c t c g t

4860 725 730 735 740 IANLAGSPAIKKGILQTVKV Cas9 725 730 735 740 IANLAGSPAIKKGILQTVKV Cas9 IANLAGSPAIKKGILQTVKV MALRAPLGAMLFPMRCVTFT NGIQGAAGGNLLADQLCHLH c a t t g c c a a t c t g g c c g g c a g c c c c g c c a t t a a g a a g g g c a t c c t g c a g a c a g t g a a g g t g t a a c g g t t a g a c c g g c c g t c g g g g c g g t a a t t c t t c c c g t a g g a c g t c t g t c a c t t c c a

4920 745 750 755 760 VDELVKVMGRHKPENIVIEM Cas9 745 750 755 760 VDELVKVMGRHKPENIVIEM Cas9 VDELVKVMGRHKPENIVIEM TSSSTFTIPRCLGSFMTISI HVLEHFHHAPVLGLVDHDFH g g t g g a c g a g c t c g t g a a a g t g a t g g g c c g g c a c a a g c c c g a g a a c a t c g t g a t c g a a a t c c a c c t g c t c g a g c a c t t t c a c t a c c c g g c c g t g t t c g g g c t c t t g t a g c a c t a g c t t t a

4980 765 770 775 780 ARENQTTQKGQKNSRERMKR Cas9 765 770 775 780 ARENQTTQKGQKNSRERMKR Cas9 ARENQTTQKGQKNSRERMKR ALSFWVVWFPCFFLRSLIFR GSLVLGGLLSLLVAALSHLP g g c c a g a g a g a a c c a g a c c a c c c a g a a g g g a c a g a a g a a c a g c c g c g a g a g a a t g a a g c g c c g g t c t c t c t t g g t c t g g t g g g t c t t c c c t g t c t t c t t g t c g g c g c t c t c t t a c t t c g c

5040 785 790 795 800 IEEGIKELGSQILKEHPVEN Cas9 785 790 795 800 IEEGIKELGSQILKEHPVEN Cas9 IEEGIKELGSQILKEHPVEN ISSPM DFLADFLQAALDQFFVGHFV g a t c g a a g a g g g c a t c a a a g a g c t g g g c a g c c a g a t c c t g a a a g a a c a c c c c g t g g a a a a c t a g c t t c t c c c g t a g t t t c t c g a c c c g t c g g t c t a g g a c t t t c t t g t g g g g c a c c t t t t

5100 805 810 815 820 TQLQNEKLYLYYLQNGRDMY Cas9 805 810 815 820 TQLQNEKLYLYYLQNGRDMY Cas9 TQLQNEKLYLYYLQNGRDMY GLQLVLLQVQVVQLIPPIHV c a c c c a g c t g c a g a a c g a g a a g c t g t a c c t g t a c t a c c t g c a g a a t g g g c g g g a t a t g t a g t g g g t c g a c g t c t t g c t c t t c g a c a t g g a c a t g a t g g a c g t c t t a c c c g c c c t a t a c a t

5160 825 830 835 840 VDQELDINRLSDYDVDHIVP Cas9 825 830 835 840 VDQELDINRLSDYDVDHIVP Cas9 VDQELDINRLSDYDVDHIVP HVLFQVDVPQGVVIHVM c g t g g a c c a g g a a c t g g a c a t c a a c c g g c t g t c c g a c t a c g a t g t g g a c c a t a t c g t g c c g c a c c t g g t c c t t g a c c t g t a g t t g g c c g a c a g g c t g a t g c t a c a c c t g g t a t a g c a c g g

5220 845 850 855 860 QSFLKDDSIDNKVLTRSDKN Cas9 845 850 855 860 QSFLKDDSIDNKVLTRSDKN Cas9 QSFLKDDSIDNKVLTRSDKN t c a g a g c t t t c t g a a g g a c g a c t c c a t c g a c a a c a a g g t g c t g a c c a g a a g c g a c a a g a a a g t c t c g a a a g a c t t c c t g c t g a g g t a g c t g t t g t t c c a c g a c t g g t c t t c g c t g t t c t t

5280 865 870 875 880 RGKSDNVPSEEVVKKMKNYW Cas9 865 870 875 880 RGKSDNVPSEEVVKKMKNYW Cas9 RGKSDNVPSEEVVKKMKNYW c c g g g g c a a g a g c g a c a a c g t g c c c t c c g a a g a g g t c g t g a a g a a g a t g a a g a a c t a c t g g g c c c c g t t c t c g c t g t t g c a c g g g a g g c t t c t c c a g c a c t t c t t c t a c t t c t t g a t g a c

5340 885 890 895 900 RQLLNAKLITQRKFDNLTKA Cas9 885 890 895 900 RQLLNAKLITQRKFDNLTKA Cas9 RQLLNAKLITQRKFDNLTKA g c g g c a g c t g c t g a a c g c c a a g c t g a t t a c c c a g a g a a a g t t c g a c a a t c t g a c c a a g g c c g c c g t c g a c g a c t t g c g g t t c g a c t a a t g g g t c t c t t t c a a g c t g t t a g a c t g g t t c c g

5400 905 910 915 920 ERGGLSELDKAGFIKRQLVE Cas9 905 910 915 920 ERGGLSELDKAGFIKRQLVE Cas9 ERGGLSELDKAGFIKRQLVE c g a g a g a g g c g g c c t g a g c g a a c t g g a t a a g g c c g g c t t c a t c a a g a g a c a g c t g g t g g a g c t c t c t c c g c c g g a c t c g c t t g a c c t a t t c c g g c c g a a g t a g t t c t c t g t c g a c c a c c t

5460 925 930 935 940 TRQITKHVAQILDSRMNTKY Cas9 925 930 935 940 TRQITKHVAQILDSRMNTKY Cas9 TRQITKHVAQILDSRMNTKY a a c c c g g c a g a t c a c a a a g c a c g t g g c a c a g a t c c t g g a c t c c c g g a t g a a c a c t a a g t a t t g g g c c g t c t a g t g t t t c g t g c a c c g t g t c t a g g a c c t g a g g g c c t a c t t g t g a t t c a t

5520 945 950 955 960 DENDKLIREVKVITLKSKLV Cas9 945 950 955 960 DENDKLIREVKVITLKSKLV Cas9 DENDKLIREVKVITLKSKLV c g a c g a g a a t g a c a a g c t g a t c c g g g a a g t g a a a g t g a t c a c c c t g a a g t c c a a g c t g g t g c t g c t c t t a c t g t t c g a c t a g g c c c t t c a c t t t c a c t a g t g g g a c t t c a g g t t c g a c c a

5580 965 970 975 980 SDFRKDFQFYKVREINNYHH Cas9 965 970 975 980 SDFRKDFQFYKVREINNYHH Cas9 SDFRKDFQFYKVREINNYHH g t c c g a t t t c c g g a a g g a t t t c c a g t t t t a c a a a g t g c g c g a g a t c a a c a a c t a c c a c c a c a g g c t a a a g g c c t t c c t a a a g g t c a a a a t g t t t c a c g c g c t c t a g t t g t t g a t g g t g g t

5640 985 990 995 1000 AHDAYLNAVVGTALIKKYPK Cas9 985 990 995 1000 AHDAYLNAVVGTALIKKYPK Cas9 AHDAYLNAVVGTALIKKYPK c g c c c a c g a c g c c t a c c t g a a c g c c g t c g t g g g a a c c g c c c t g a t c a a a a a g t a c c c t a a g c g g g t g c t g c g g a t g g a c t t g c g g c a g c a c c c t t g g c g g g a c t a g t t t t t c a t g g g a t t

5700 1005 1010 1015 1020 LESEFVYGDYKVYDVRKMIA Cas9 1005 1010 1015 1020 LESEFVYGDYKVYDVRKMIA Cas9 LESEFVYGDYKVYDVRKMIA g c t g g a a a g c g a g t t c g t g t a c g g c g a c t a c a a g g t g t a c g a c g t g c g g a a g a t g a t c g c c g a c c t t t c g c t c a a g c a c a t g c c g c t g a t g t t c c a c a t g c t g c a c g c c t t c t a c t a g c g

5760 1025 1030 1035 1040 KSEQEIGKATAKYFFYSNIM Cas9 1025 1030 1035 1040 KSEQEIGKATAKYFFYSNIM Cas9 KSEQEIGKATAKYFFYSNIM c a a g a g c g a g c a g g a a a t c g g c a a g g c t a c c g c c a a g t a c t t c t t c t a c a g c a a c a t c a t g t t c t c g c t c g t c c t t t a g c c g t t c c g a t g g c g g t t c a t g a a g a a g a t g t c g t t g t a g t a

5820 1045 1050 1055 1060 NFFKTEITLANGEIRKRPLI Cas9 1045 1050 1055 1060 NFFKTEITLANGEIRKRPLI Cas9 NFFKTEITLANGEIRKRPLI g a a c t t t t t c a a g a c c g a g a t t a c c c t g g c c a a c g g c g a g a t c c g g a a g c g g c c t c t g a t c t t g a a a a a g t t c t g g c t c t a a t g g g a c c g g t t g c c g c t c t a g g c c t t c g c c g g a g a c t a

5880 1065 1070 1075 1080 ETNGETGEIVWDKGRDFATV Cas9 1065 1070 1075 1080 ETNGETGEIVWDKGRDFATV Cas9 ETNGETGEIVWDKGRDFATV c g a g a c a a a c g g c g a a a c c g g g g a g a t c g t g t g g g a t a a g g g c c g g g a t t t t g c c a c c g t g c t c t g t t t g c c g c t t t g g c c c c t c t a g c a c a c c c t a t t c c c g g c c c t a a a a c g g t g g c a

5940 1085 1090 1095 1100 RKVLSMPQVNIVKKTEVQTG Cas9 1085 1090 1095 1100 RKVLSMPQVNIVKKTEVQTG Cas9 RKVLSMPQVNIVKKTEVQTG g c g g a a a g t g c t g a g c a t g c c c c a a g t g a a t a t c g t g a a a a a g a c c g a g g t g c a g a c a g g c g c c t t t c a c g a c t c g t a c g g g g t t c a c t t a t a g c a c t t t t t c t g g c t c c a c g t c t g t c c

6000 1105 1110 1115 1120 GFSKESILPKRNSDKLIARK Cas9 1105 1110 1115 1120 GFSKESILPKRNSDKLIARK Cas9 GFSKESILPKRNSDKLIARK c g g c t t c a g c a a a g a g t c t a t c c t g c c c a a g a g g a a c a g c g a t a a g c t g a t c g c c a g a a a g c c g a a g t c g t t t c t c a g a t a g g a c g g g t t c t c c t t g t c g c t a t t c g a c t a g c g g t c t t t

6060 1125 1130 1135 1140 KDWDPKKYGGFDSPTVAYSV Cas9 1125 1130 1135 1140 KDWDPKKYGGFDSPTVAYSV Cas9 KDWDPKKYGGFDSPTVAYSV g a a g g a c t g g g a c c c t a a g a a g t a c g g c g g c t t c g a c a g c c c c a c c g t g g c c t a t t c t g t c t t c c t g a c c c t g g g a t t c t t c a t g c c g c c g a a g c t g t c g g g g t g g c a c c g g a t a a g a c a

6120 1145 1150 1155 1160 LVVAKVEKGKSKKLKSVKEL Cas9 1145 1150 1155 1160 LVVAKVEKGKSKKLKSVKEL Cas9 LVVAKVEKGKSKKLKSVKEL g c t g g t g g t g g c c a a a g t g g a a a a g g g c a a g t c c a a g a a a c t g a a g a g t g t g a a a g a g c t c g a c c a c c a c c g g t t t c a c c t t t t c c c g t t c a g g t t c t t t g a c t t c t c a c a c t t t c t c g a

BsaBI* 6180 1165 1170 1175 1180 LGITIMERSSFEKNPIDFLE Cas9 1165 1170 1175 1180 LGITIMERSSFEKNPIDFLE Cas9 LGITIMERSSFEKNPIDFLE g c t g g g g a t c a c c a t c a t g g a a a g a a g c a g c t t c g a g a a g a a t c c c a t c g a c t t t c t g g a c g a c c c c t a g t g g t a g t a c c t t t c t t c g t c g a a g c t c t t c t t a g g g t a g c t g a a a g a c c t

6240 1185 1190 1195 1200 AKGYKEVKKDLIIKLPKYSL Cas9 1185 1190 1195 1200 AKGYKEVKKDLIIKLPKYSL Cas9 AKGYKEVKKDLIIKLPKYSL a g c c a a g g g c t a c a a a g a a g t g a a a a a g g a c c t g a t c a t c a a g c t g c c t a a g t a c t c c c t t c g g t t c c c g a t g t t t c t t c a c t t t t t c c t g g a c t a g t a g t t c g a c g g a t t c a t g a g g g a

6300 1205 1210 1215 1220 FELENGRKRMLASAGELQKG Cas9 1205 1210 1215 1220 FELENGRKRMLASAGELQKG Cas9 FELENGRKRMLASAGELQKG g t t c g a g c t g g a a a a c g g c c g g a a g a g a a t g c t g g c c t c t g c c g g c g a a c t g c a g a a g g g c a a g c t c g a c c t t t t g c c g g c c t t c t c t t a c g a c c g g a g a c g g c c g c t t g a c g t c t t c c c

6360 1225 1230 1235 1240 NELALPSKYVNFLYLASHYE Cas9 1225 1230 1235 1240 NELALPSKYVNFLYLASHYE Cas9 NELALPSKYVNFLYLASHYE a a a c g a a c t g g c c c t g c c c t c c a a a t a t g t g a a c t t c c t g t a c c t g g c c a g c c a c t a t g a t t t g c t t g a c c g g g a c g g g a g g t t t a t a c a c t t g a a g g a c a t g g a c c g g t c g g t g a t a c t

6420 1245 1250 1255 1260 KLKGSPEDNEQKQLFVEQHK Cas9 1245 1250 1255 1260 KLKGSPEDNEQKQLFVEQHK Cas9 KLKGSPEDNEQKQLFVEQHK g a a g c t g a a g g g c t c c c c c g a g g a t a a t g a g c a g a a a c a g c t g t t t g t g g a a c a g c a c a a c t t c g a c t t c c c g a g g g g g c t c c t a t t a c t c g t c t t t g t c g a c a a a c a c c t t g t c g t g t t

6480 1265 1270 1275 1280 HYLDEIIEQISEFSKRVILA Cas9 1265 1270 1275 1280 HYLDEIIEQISEFSKRVILA Cas9 HYLDEIIEQISEFSKRVILA g c a c t a c c t g g a c g a g a t c a t c g a g c a g a t c a g c g a g t t c t c c a a g a g a g t g a t c c t g g c c g t g a t g g a c c t g c t c t a g t a g c t c g t c t a g t c g c t c a a g a g g t t c t c t c a c t a g g a c c g

6540 1285 1290 1295 1300 DANLDKVLSAYNKHRDKPIR Cas9 1285 1290 1295 1300 DANLDKVLSAYNKHRDKPIR Cas9 DANLDKVLSAYNKHRDKPIR c g a c g c t a a t c t g g a c a a a g t g c t g t c c g c c t a c a a c a a g c a c c g g g a t a a g c c c a t c a g g c t g c g a t t a g a c c t g t t t c a c g a c a g g c g g a t g t t g t t c g t g g c c c t a t t c g g g t a g t c

6600 1305 1310 1315 1320 EQAENIIHLFTLTNLGAPAA Cas9 1305 1310 1315 1320 EQAENIIHLFTLTNLGAPAA Cas9 EQAENIIHLFTLTNLGAPAA a g a g c a g g c c g a g a a t a t c a t c c a c c t g t t t a c c c t g a c c a a t c t g g g a g c c c c t g c c g c t c t c g t c c g g c t c t t a t a g t a g g t g g a c a a a t g g g a c t g g t t a g a c c c t c g g g g a c g g c g

6660 1325 1330 1335 1340 FKYFDTTIDRKRYTSTKEVL Cas9 1325 1330 1335 1340 FKYFDTTIDRKRYTSTKEVL Cas9 FKYFDTTIDRKRYTSTKEVL c t t c a a g t a c t t t g a c a c c a c c a t c g a c c g g a a g a g g t a c a c c a g c a c c a a a g a g g t g c t g a a g t t c a t g a a a c t g t g g t g g t a g c t g g c c t t c t c c a t g t g g t c g t g g t t t c t c c a c g a

6720 1345 1350 1355 1360 DATLIHQSITGLYETRIDLS Cas9 1345 1350 1355 1360 DATLIHQSITGLYETRIDLS Cas9 DATLIHQSITGLYETRIDLS g g a c g c c a c c c t g a t c c a c c a g a g c a t c a c c g g c c t g t a c g a g a c a c g g a t c g a c c t g t c c c t g c g g t g g g a c t a g g t g g t c t c g t a g t g g c c g g a c a t g c t c t g t g c c t a g c t g g a c a g

6780 1365 QLGGD (in frame with Cas9) KRPAATKKAGQAKKK Cas9 1365 QLGGD 1 5 10 15 KRPAATKKAGQAKKK Cas9 nucleoplasmin NLS QLGGDKRPAATKKAGQAKKK t c a g c t g g g a g g c g a c a a g c g a c c t g c c g c c a c a a a g a a g g c t g g a c a g g c t a a g a a g a a a g t c g a c c c t c c g c t g t t c g c t g g a c g g c g g t g t t t c t t c c g a c c t g t c c g a t t c t t c t t

BamHI 6840 (in frame with Cas9) KDYKDDDDKGSGATNFSLLK K 1 5 DYKDDDDK GSG 1 5 ATNFSLLK nucleoplasmin NLS FLAG P2A KDYKDDDDKGSGATNFSLLK g a a a g a t t a c a a a g a c g a t g a c g a t a a g g g a t c c g g c g c a a c a a a c t t c t c t c t g c t g a a c t t t c t a a t g t t t c t g c t a c t g c t a t t c c c t a g g c c g c g t t g t t t g a a g a g a g a c g a c t t

6900 (in frame with Cas9) QAGDVEENPGPTEYKPTVRL 10 15 QAGDVEENPGP 5 10 TEYKPTVRL P2A PuroR QAGDVEENPGPTEYKPTVRL a c a a g c c g g a g a t g t c g a a g a g a a t c c t g g a c c g a c c g a g t a c a a g c c c a c g g t g c g c c t t g t t c g g c c t c t a c a g c t t c t c t t a g g a c c t g g c t g g c t c a t g t t c g g g t g c c a c g c g g a

BsiWITth111IPflFI 6960 (in frame with Cas9) ATRDDVPRAVRTLAAAFADY 15 20 25 30 ATRDDVPRAVRTLAAAFADY PuroR ATRDDVPRAVRTLAAAFADY c g c c a c c c g c g a c g a c g t c c c c a g g g c c g t a c g c a c c c t c g c c g c c g c g t t c g c c g a c t a g c g g t g g g c g c t g c t g c a g g g g t c c c g g c a t g c g t g g g a g c g g c g g c g c a a g c g g c t g a t

BstEIIRsrII 7020 (in frame with Cas9) PATRHTVDPDRHIERVTELQ 35 40 45 50 PATRHTVDPDRHIERVTELQ PuroR PATRHTVDPDRHIERVTELQ c c c c g c c a c g c g c c a c a c c g t c g a t c c g g a c c g c c a c a t c g a g c g g g t c a c c g a g c t g c a g g g g c g g t g c g c g g t g t g g c a g c t a g g c c t g g c g g t g t a g c t c g c c c a g t g g c t c g a c g t

7080 (in frame with Cas9) ELFLTRVGLDIGKVWVADDG 55 60 65 70 ELFLTRVGLDIGKVWVADDG PuroR ELFLTRVGLDIGKVWVADDG a g a a c t c t t c c t c a c g c g c g t c g g g c t c g a c a t c g g c a a g g t g t g g g t c g c g g a c g a c g g t c t t g a g a a g g a g t g c g c g c a g c c c g a g c t g t a g c c g t t c c a c a c c c a g c g c c t g c t g c c

7140 (in frame with Cas9) AAVAVWTTPESVEAGAVFAE 75 80 85 90 AAVAVWTTPESVEAGAVFAE PuroR AAVAVWTTPESVEAGAVFAE c g c c g c g g t g g c g g t c t g g a c c a c g c c g g a g a g c g t c g a a g c g g g g g c g g t g t t c g c c g a g c g g c g c c a c c g c c a g a c c t g g t g c g g c c t c t c g c a g c t t c g c c c c c g c c a c a a g c g g c t

7200 (in frame with Cas9) IGPRMAELSGSRLAAQQQME 95 100 105 110 IGPRMAELSGSRLAAQQQME PuroR IGPRMAELSGSRLAAQQQME g a t c g g c c c g c g c a t g g c c g a g t t g a g c g g t t c c c g g c t g g c c g c g c a g c a a c a g a t g g a c t a g c c g g g c g c g t a c c g g c t c a a c t c g c c a a g g g c c g a c c g g c g c g t c g t t g t c t a c c t

7260 (in frame with Cas9) GLLAPHRPKEPAWFLATVGV 115 120 125 130 GLLAPHRPKEPAWFLATVGV PuroR GLLAPHRPKEPAWFLATVGV a g g c c t c c t g g c g c c g c a c c g g c c c a a g g a g c c c g c g t g g t t c c t g g c c a c c g t c g g a g t t c c g g a g g a c c g c g g c g t g g c c g g g t t c c t c g g g c g c a c c a a g g a c c g g t g g c a g c c t c a

7320 (in frame with Cas9) SPDHQGKGLGSAVVLPGVEA 135 140 145 150 SPDHQGKGLGSAVVLPGVEA PuroR SPDHQGKGLGSAVVLPGVEA c t c g c c c g a c c a c c a g g g c a a g g g t c t g g g c a g c g c c g t c g t g c t c c c c g g a g t g g a g g c g a g c g g g c t g g t g g t c c c g t t c c c a g a c c c g t c g c g g c a g c a c g a g g g g c c t c a c c t c c g

7380 (in frame with Cas9) AERAGVPAFLETSAPRNLPF 155 160 165 170 AERAGVPAFLETSAPRNLPF PuroR G C A A C C T C C C C T T Puro-F AERAGVPAFLETSAPRNLPF g g c c g a g c g c g c c g g g g t g c c c g c c t t c c t g g a g a c c t c c g c g c c c c g c a a c c t c c c c t t c c g g c t c g c g c g g c c c c a c g g g c g g a a g g a c c t c t g g a g g c g c g g g g c g t t g g a g g g g a a

7440 (in frame with Cas9) YERLGFTVTADVEVPEGPRT 175 180 185 190 YERLGFTVTADVEVPEGPRT PuroR C T A C G A G C Puro-F YERLGFTVTADVEVPEGPRT c t a c g a g c g g c t c g g c t t c a c c g t c a c c g c c g a c g t c g a g g t g c c c g a a g g a c c g c g c a c g a t g c t c g c c g a g c c g a a g t g g c a g t g g c g g c t g c a g c t c c a c g g g c t t c c t g g c g c g t g

7500 (in frame with Cas9) WCMTRKPGA* WPRE 195 200 WCMTRKPGA* PuroR WCMTRKPGA* c t g g t g c a t g a c c c g c a a g c c c g g t g c c t g a a c g c g t t a a g t c g a c a a t c a a c c t c t g g a g a c c a c g t a c t g g g c g t t c g g g c c a c g g a c t t g c g c a a t t c a g c t g t t a g t t g g a g a c c t

7560 WPRE A C A A C G A G G A A A A T G C G A T A C WPRE-R t t a c a a a a t t t g t g a a a g a t t g a c t g g t a t t c t t a a c t a t g t t g c t c c t t t t a c g c t a t g a a t g t t t t a a a c a c t t t c t a a c t g a c c a t a a g a a t t g a t a c a a c g a g g a a a a t g c g a t a c

7620 WPRE MPLYHAIASRMAFIF MLLLPVWLSF t g g a t a c g c t g c t t t a a t g c c t t t g t a t c a t g c t a t t g c t t c c c g t a t g g c t t t c a t t t t a c c t a t g c g a c g a a a t t a c g g a a a c a t a g t a c g a t a a c g a a g g g c a t a c c g a a a g t a a a a

7680 WPRE SSLYKSWLLSLYEELWPVVR SPPCINPGCCLFMRSCGPLS c t c c t c c t t g t a t a a a t c c t g g t t g c t g t c t c t t t a t g a g g a g t t g t g g c c c g t t g t c a g g a g g a g g a a c a t a t t t a g g a c c a a c g a c a g a g a a a t a c t c c t c a a c a c c g g g c a a c a g t c

7740 WPRE QRGVVCTVFADATPTGWGIA GNVAWCALCLLTQPPLVGAL g c a a c g t g g c g t g g t g t g c a c t g t g t t t g c t g a c g c a a c c c c c a c t g g t t g g g g c a t t g c c g t t g c a c c g c a c c a c a c g t g a c a c a a a c g a c t g c g t t g g g g g t g a c c a a c c c c g t a a c g

7800 WPRE TTCQLLSGTFAFPLPIATAE PPPVSSFPGLSLSPSLLPRR c a c c a c c t g t c a g c t c c t t t c c g g g a c t t t c g c t t t c c c c c t c c c t a t t g c c a c g g c g g a g t g g t g g a c a g t c g a g g a a a g g c c c t g a a a g c g a a a g g g g g a g g g a t a a c g g t g c c g c c t

7860 WPRE LIAACLARCWTGARLLGTDN NSSPPALPAAGQGLGCWALT a c t c a t c g c c g c c t g c c t t g c c c g c t g c t g g a c a g g g g c t c g g c t g t t g g g c a c t g a c a a t g a g t a g c g g c g g a c g g a a c g g g c g a c g a c c t g t c c c c g a g c c g a c a a c c c g t g a c t g t t

7920 WPRE (in frame with Factor Xa site) *RGKRPQEGTNGG SVVLSGKSSSFPWLLACVAT IPWCCRGNHRPFLGCSPVLP t t c c g t g g t g t t g t c g g g g a a a t c a t c g t c c t t t c c t t g g c t g c t c g c c t g t g t t g c c a c a a g g c a c c a c a a c a g c c c c t t t a g t a g c a g g a a a g g a a c c g a c g a g c g g a c a c a a c g g t g

7980 WPRE (in frame with Factor Xa site) PNQAPRGEAVDR 1 RGEI Factor Xa site WILRGTSFCYVPSALNPADL PGFCAGRPSATSLRPSIQRT c t g g a t t c t g c g c g g g a c g t c c t t c t g c t a c g t c c c t t c g g c c c t c a a t c c a g c g g a c c t g a c c t a a g a c g c g c c c t g c a g g a a g a c g a t g c a g g g a a g c c g g g a g t t a g g t c g c c t g g a

8040 WPRE PSRGLLPALRPLPRLRLRPQ FLPAACCRLCGLFRVFAFAL t c c t t c c c g c g g c c t g c t g c c g g c t c t g c g g c c t c t t c c g c g t c t t c g c c t t c g c c c t c a a g g a a g g g c g c c g g a c g a c g g c c g a g a c g c c g g a g a a g g c g c a g a a g c g g a a g c g g g a g t

8100 WPRE TSRISLWAASPRRL* RRVGSPFGPPPRVDFKTNDL g a c g a g t c g g a t c t c c c t t t g g g c c g c c t c c c c g c g t c g a c t t t a a g a c c a a t g a c t t a c c t g c t c a g c c t a g a g g g a a a c c c g g c g g a g g g g c g c a g c t g a a a t t c t g g t t a c t g a a t g

8160 3' LTR ( Δ U3) QGSCRS* a a g g c a g c t g t a g a t c t t a g c c a c t t t t t a a a a g a a a a g g g g g g a c t g g a a g g g c t a a t t t t c c g t c g a c a t c t a g a a t c g g t g a a a a a t t t t c t t t t c c c c c c t g a c c t t c c c g a t t a a

8220 3' LTR ( Δ U3) c a c t c c c a a c g a a g a c a a g a t c t g c t t t t t g c t t g t a c t g g g t c t c t c t g g t t a g a c c a g g t g a g g g t t g c t t c t g t t c t a g a c g a a a a a c g a a c a t g a c c c a g a g a g a c c a a t c t g g t c

8280 3' LTR ( Δ U3) a t c t g a g c c t g g g a g c t c t c t g g c t a a c t a g g g a a c c c a c t g c t t a a g c c t c a a t a a a g c t a g a c t c g g a c c c t c g a g a g a c c g a t t g a t c c c t t g g g t g a c g a a t t c g g a g t t a t t t c g

8340 3' LTR ( Δ U3) t t g c c t t g a g t g c t t c a a g t a g t g t g t g c c c g t c t g t t g t g t g a c t c t g g t a a c t a g a g a a a c g g a a c t c a c g a a g t t c a t c a c a c a c g g g c a g a c a a c a c a c t g a g a c c a t t g a t c t c t

PmeI 8400 3' LTR ( Δ U3) t c c c t c a g a c c c t t t t a g t c a g t g t g g a a a a t c t c t a g c a g g g c c c g t t t a a a c c c g c t g a g g g a g t c t g g g a a a a t c a g t c a c a c c t t t t a g a g a t c g t c c c g g g c a a a t t t g g g c g a c

8460 bGH poly(A) signal G G A G C T G A C A C G G A A G A T BGH-rev a t c a g c c t c g a c t g t g c c t t c t a g t t g c c a g c c a t c t g t t g t t t g c c c c t c c c c c g t g c c t a g t c g g a g c t g a c a c g g a a g a t c a a c g g t c g g t a g a c a a c a a a c g g g g a g g g g g c a c g g

8520 bGH poly(A) signal t t c c t t g a c c c t g g a a g g t g c c a c t c c c a c t g t c c t t t c c t a a t a a a a t g a g g a a a t t g c a a g g a a c t g g g a c c t t c c a c g g t g a g g g t g a c a g g a a a g g a t t a t t t t a c t c c t t t a a c g

8580 bGH poly(A) signal a t c g c a t t g t c t g a g t a g g t g t c a t t c t a t t c t g g g g g g t g g g g t g g g g c a g g a c a g c a a t a g c g t a a c a g a c t c a t c c a c a g t a a g a t a a g a c c c c c c a c c c c a c c c c g t c c t g t c g t t

8640 bGH poly(A) signal g g g g g a g g a t t g g g a a g a c a a t a g c a g g c a t g c t g g g g a t g c g g t g g g c t c t a t g g c t t c c c c c c t c c t a a c c c t t c t g t t a t c g t c c g t a c g a c c c c t a c g c c a c c c g a g a t a c c g a a g

8700 f1 ori t g a g g c g g a a a g a a c c a g c t g g g g c t c t a g g g g g t a t c c c c a c g c g c c c t g t a g c g g c g c a c t c c g c c t t t c t t g g t c g a c c c c g a g a t c c c c c a t a g g g g t g c g c g g g a c a t c g c c g c g

8760 f1 ori a t t a a g c g c g g c g g g t g t g g t g g t t a c g c g c a g c g t g a c c g c t a c a c t t g c c a g c g c c c t t a a t t c g c g c c g c c c a c a c c a c c a a t g c g c g t c g c a c t g g c g a t g t g a a c g g t c g c g g g a

8820 f1 ori G A G G A A A G C G A A A G A A G G G A F1ori-R a g c g c c c g c t c c t t t c g c t t t c t t c c c t t c c t t t c t c g c c a c g t t c g c c g g c t t t c c c c g t c g c g g g c g a g g a a a g c g a a a g a a g g g a a g g a a a g a g c g g t g c a a g c g g c c g a a a g g g g c

8880 f1 ori t c a a g c t c t a a a t c g g g g g c t c c c t t t a g g g t t c c g a t t t a g t g c t t t a c g g c a c c t c g a a g t t c g a g a t t t a g c c c c c g a g g g a a a t c c c a a g g c t a a a t c a c g a a a t g c c g t g g a g c t

8940 f1 ori c c c c a a a a a a c t t g a t t a g g g t g a t g g t t c a c g t a g t g g g c c a t c g c c c t g a t a g a c g g t g g g g t t t t t t g a a c t a a t c c c a c t a c c a a g t g c a t c a c c c g g t a g c g g g a c t a t c t g c c a

9000 f1 ori G T G G A C T C T T G T T C C A A A C T G G F1ori-F t t t t c g c c c t t t g a c g t t g g a g t c c a c g t t c t t t a a t a g t g g a c t c t t g t t c c a a a c t g g a a a a g c g g g a a a c t g c a a c c t c a g g t g c a a g a a a t t a t c a c c t g a g a a c a a g g t t t g a c c

9060 f1 ori a a c a a c a c t c a a c c c t a t c t c g g t c t a t t c t t t t g a t t t a t a a g g g a t t t t g c c g a t t t c t t g t t g t g a g t t g g g a t a g a g c c a g a t a a g a a a a c t a a a t a t t c c c t a a a a c g g c t a a a g

9120 f1 ori C C pBABE 3' g g c c t a t t g g t t a a a a a a t g a g c t g a t t t a a c a a a a a t t t a a c g c g a a t t a a t t c t g t g g c c g g a t a a c c a a t t t t t t a c t c g a c t a a a t t g t t t t t a a a t t g c g c t t a a t t a a g a c a c c

9180 SV40 promoter T T A C A C A C A G T C A A T C C C A pBABE 3' a a t g t g t g t c a g t t a g g g t g t g g a a a g t c c c c a g g c t c c c c a g c a g g c a g a a g t a t g c a a t t a c a c a c a g t c a a t c c c a c a c c t t t c a g g g g t c c g a g g g g t c g t c c g t c t t c a t a c g t t

9240 SV40 promoter a g c a t g c a t c t c a a t t a g t c a g c a a c c a g g t g t g g a a a g t c c c c a g g c t c c c c a g c a g g c t c g t a c g t a g a g t t a a t c a g t c g t t g g t c c a c a c c t t t c a g g g g t c c g a g g g g t c g t c c g

9300 SV40 promoter a g a a g t a t g c a a a g c a t g c a t c t c a a t t a g t c a g c a a c c a t a g t c c c g c c c c t a a c t c c g t c t t c a t a c g t t t c g t a c g t a g a g t t a a t c a g t c g t t g g t a t c a g g g c g g g g a t t g a g g c

9360 SV40 promoter SV40 ori c c c a t c c c g c c c c t a a c t c c g c c c a g t t c c g c c c a t t c t c c g c c c c a t g g c t g a c t a a t t g g g t a g g g c g g g g a t t g a g g c g g g t c a a g g c g g g t a a g a g g c g g g g t a c c g a c t g a t t a a

9420 SV40 promoter SV40 ori T A T T T A T G C A G A G G C C G A G G SV40pro-F t t t t t t a t t t a t g c a g a g g c c g a g g c c g c c t c t g c c t c t g a g c t a t t c c a g a a g t a g t g a a a a a a a t a a a t a c g t c t c c g g c t c c g g c g g a g a c g g a g a c t c g a t a a g g t c t t c a t c a c t

AvrII 9480 SV40 promoter SV40 ori g g a g g c t t t t t t g g a g g c c t a g g c t t t t g c a a a a a g c t c c c g g g a g c t t g t a t a t c c a t t c c t c c g a a a a a a c c t c c g g a t c c g a a a a c g t t t t t c g a g g g c c c t c g a a c a t a t a g g t a a

9540 EM7 promoter t t c g g a t c t g a t c a g c a c g t g t t g a c a a t t a a t c a t c g g c a t a g t a t a t c g g c a t a g t a t a a g c c t a g a c t a g t c g t g c a c a a c t g t t a a t t a g t a g c c g t a t c a t a t a g c c g t a t c a t a

9600 1 5 10 MAKLTSAVPVL EM7 promoter BleoR a a t a c g a c a a g g t g a g g a a c t a a a c c a t g g c c a a g t t g a c c a g t g c c g t t c c g g t g c t c a t t a t g c t g t t c c a c t c c t t g a t t t g g t a c c g g t t c a a c t g g t c a c g g c a a g g c c a c g a g t

MauBI 9660 15 20 25 30 TARDVAGAVEFWTDRLGFSR BleoR c c g c g c g c g a c g t c g c c g g a g c g g t c g a g t t c t g g a c c g a c c g g c t c g g g t t c t c c c g g g g g c g c g c g c t g c a g c g g c c t c g c c a g c t c a a g a c c t g g c t g g c c g a g c c c a a g a g g g c c c

SgrAI 9720 35 40 45 50 DFVEDDFAGVVRDDVTLFIS BleoR a c t t c g t g g a g g a c g a c t t c g c c g g t g t g g t c c g g g a c g a c g t g a c c c t g t t c a t c a g c g t g a a g c a c c t c c t g c t g a a g c g g c c a c a c c a g g c c c t g c t g c a c t g g g a c a a g t a g t c g c

9780 55 60 65 70 AVQDQVVPDNTLAWVWVRGL BleoR c g g t c c a g g a c c a g g t g g t g c c g g a c a a c a c c c t g g c c t g g g t g t g g g t g c g c g g c c t g g g c c a g g t c c t g g t c c a c c a c g g c c t g t t g t g g g a c c g g a c c c a c a c c c a c g c g c c g g a c c

9840 75 80 85 90 DELYAEWSEVVSTNFRDASG BleoR a c g a g c t g t a c g c c g a g t g g t c g g a g g t c g t g t c c a c g a a c t t c c g g g a c g c c t c c g g g c t g c t c g a c a t g c g g c t c a c c a g c c t c c a g c a c a g g t g c t t g a a g g c c c t g c g g a g g c c c g

FseI 9900 95 100 105 110 PAMTEIGEQPWGREFALRDP BleoR c g g c c a t g a c c g a g a t c g g c g a g c a g c c g t g g g g g c g g g a g t t c g c c c t g c g c g a c c c g g g c c g g t a c t g g c t c t a g c c g c t c g t c g g c a c c c c c g c c c t c a a g c g g g a c g c g c t g g g c c

9960 115 120 125 AGNCVHFVAEEQD* BleoR c c g g c a a c t g c g t g c a c t t c g t g g c c g a g g a g c a g g a c t g a c a c g t g c t a c g a g a t t t c g g g c c g t t g a c g c a c g t g a a g c a c c g g c t c c t c g t c c t g a c t g t g c a c g a t g c t c t a a a g c

10,020 a t t c c a c c g c c g c c t t c t a t g a a a g g t t g g g c t t c g g a a t c g t t t t c c g g g a c g c c g g c t t a a g g t g g c g g c g g a a g a t a c t t t c c a a c c c g a a g c c t t a g c a a a a g g c c c t g c g g c c g a

10,080 SV40 poly(A) signal g g a t g a t c c t c c a g c g c g g g g a t c t c a t g c t g g a g t t c t t c g c c c a c c c c a a c t t g t t t a c c t a c t a g g a g g t c g c g c c c c t a g a g t a c g a c c t c a a g a a g c g g g t g g g g t t g a a c a a a t

10,140 SV40 poly(A) signal C G T T A T C G T A G T G T T T A A A G SV40pA-R t t g c a g c t t a t a a t g g t t a c a a a t a a a g c a a t a g c a t c a c a a a t t t c a c a a a t a a a g c a t a a c g t c g a a t a t t a c c a a t g t t t a t t t c g t t a t c g t a g t g t t t a a a g t g t t t a t t t c g t a

10,200 SV40 poly(A) signal G T G G T T T G T C C A A A C T C A T C EBV-rev t t t t t t c a c t g c a t t c t a g t t g t g g t t t g t c c a a a c t c a t c a a t g t a t c t t a t c a t g t c t a a a a a a g t g a c g t a a g a t c a a c a c c a a a c a g g t t t g a g t a g t t a c a t a g a a t a g t a c a g a

BstZ17I 10,260 M13 rev C A G T A T C G A C A A A G G A C M13 Reverse G G A C A C A M13/pUC Reverse g t a t a c c g t c g a c c t c t a g c t a g a g c t t g g c g t a a t c a t g g t c a t a g c t g t t t c c t g t g t c a t a t g g c a g c t g g a g a t c g a t c t c g a a c c g c a t t a g t a c c a g t a t c g a c a a a g g a c a c a

10,320 -10 -35 lac operator lac promoter C T T T A A C A A T A G G C G A M13/pUC Reverse g a a a t t g t t a t c c g c t c a c a a t t c c a c a c a a c a t a c g a g c c g g a a g c a t a a a g t g t a a a g c t t t a a c a a t a g g c g a g t g t t a a g g t g t g t t g t a t g c t c g g c c t t c g t a t t t c a c a t t t c

10,380 CAP binding site c c t g g g g t g c c t a a t g a g t g a g c t a a c t c a c a t t a a t t g c g t t g c g c t c a c t g c c c g c t t g g a c c c c a c g g a t t a c t c a c t c g a t t g a g t g t a a t t a a c g c a a c g c g a g t g a c g g g c g a a

10,440 t c c a g t c g g g a a a c c t g t c g t g c c a g c t g c a t t a a t g a a t c g g c c a a c g c g c g g g g a g a g a g g t c a g c c c t t t g g a c a g c a c g g t c g a c g t a a t t a c t t a g c c g g t t g c g c g c c c c t c t c

10,500 G A G C G A G T G A C T G A G C G A L4440 g c g g t t t g c g t a t t g g g c g c t c t t c c g c t t c c t c g c t c a c t g a c t c g c t g c g c t c g g t c g c g c c a a a c g c a t a a c c c g c g a g a a g g c g a a g g a g c g a g t g a c t g a g c g a c g c g a g c c a g c

10,560 t t c g g c t g c g g c g a g c g g t a t c a g c t c a c t c a a a g g c g g t a a t a c g g t t a t c c a c a g a a t a a g c c g a c g c c g c t c g c c a t a g t c g a g t g a g t t t c c g c c a t t a t g c c a a t a g g t g t c t t a

PciI 10,620 c a g g g g a t a a c g c a g g a a a g a a c a t g t g a g c a a a a g g c c a g c a a a a g g c c a g g a a c c g t a g t c c c c t a t t g c g t c c t t t c t t g t a c a c t c g t t t t c c g g t c g t t t t c c g g t c c t t g g c a t

10,680 ori a a a a g g c c g c g t t g c t g g c g t t t t t c c a t a g g c t c c g c c c c c c t g a c g a g c a t c a c a a a a t t t t c c g g c g c a a c g a c c g c a a a a a g g t a t c c g a g g c g g g g g g a c t g c t c g t a g t g t t t t

10,740 ori T T T C T A T G G T C C G C A A A G pBR322ori-F a t c g a c g c t c a a g t c a g a g g t g g c g a a a c c c g a c a g g a c t a t a a a g a t a c c a g g c g t t t c t a g c t g c g a g t t c a g t c t c c a c c g c t t t g g g c t g t c c t g a t a t t t c t a t g g t c c g c a a a g

10,800 ori G G pBR322ori-F c c c c t g g a a g c t c c c t c g t g c g c t c t c c t g t t c c g a c c c t g c c g c t t a c c g g a t a c c t g t g g g g a c c t t c g a g g g a g c a c g c g a g a g g a c a a g g c t g g g a c g g c g a a t g g c c t a t g g a c a

10,860 ori c c g c c t t t c t c c c t t c g g g a a g c g t g g c g c t t t c t c a t a g c t c a c g c t g t a g g t a t c t c a g g c g g a a a g a g g g a a g c c c t t c g c a c c g c g a a a g a g t a t c g a g t g c g a c a t c c a t a g a g t

10,920 ori g t t c g g t g t a g g t c g t t c g c t c c a a g c t g g g c t g t g t g c a c g a a c c c c c c g t t c a g c c c g c a a g c c a c a t c c a g c a a g c g a g g t t c g a c c c g a c a c a c g t g c t t g g g g g g c a a g t c g g g c

10,980 ori a c c g c t g c g c c t t a t c c g g t a a c t a t c g t c t t g a g t c c a a c c c g g t a a g a c a c g a c t t a t t g g c g a c g c g g a a t a g g c c a t t g a t a g c a g a a c t c a g g t t g g g c c a t t c t g t g c t g a a t a

11,040 ori c g c c a c t g g c a g c a g c c a c t g g t a a c a g g a t t a g c a g a g c g a g g t a t g t a g g c g g t g c t a g c g g t g a c c g t c g t c g g t g a c c a t t g t c c t a a t c g t c t c g c t c c a t a c a t c c g c c a c g a t

11,100 ori c a g a g t t c t t g a a g t g g t g g c c t a a c t a c g g c t a c a c t a g a a g a a c a g t a t t t g g t a t c t g t c t c a a g a a c t t c a c c a c c g g a t t g a t g c c g a t g t g a t c t t c t t g t c a t a a a c c a t a g a

11,160 ori g c g c t c t g c t g a a g c c a g t t a c c t t c g g a a a a a g a g t t g g t a g c t c t t g a t c c g g c a a a c c g c g a g a c g a c t t c g g t c a a t g g a a g c c t t t t t c t c a a c c a t c g a g a a c t a g g c c g t t t g

11,220 ori a a a c c a c c g c t g g t a g c g g t g g t t t t t t t g t t t g c a a g c a g c a g a t t a c g c g c a g a a a a a t t t g g t g g c g a c c a t c g c c a c c a a a a a a a c a a a c g t t c g t c g t c t a a t g c g c g t c t t t t t

11,280 ori a a g g a t c t c a a g a a g a t c c t t t g a t c t t t t c t a c g g g g t c t g a c g c t c a g t g g a a c g a a a t t c c t a g a g t t c t t c t a g g a a a c t a g a a a a g a t g c c c c a g a c t g c g a g t c a c c t t g c t t t

11,340 a c t c a c g t t a a g g g a t t t t g g t c a t g a g a t t a t c a a a a a g g a t c t t c a c c t a g a t c c t t t t g a g t g c a a t t c c c t a a a a c c a g t a c t c t a a t a g t t t t t c c t a g a a g t g g a t c t a g g a a a

11,400 t a a a t t a a a a a t g a a g t t t t a a a t c a a t c t a a a g t a t a t a t g a g t a a a c t t g g t c t g a c a a t t t a a t t t t t a c t t c a a a a t t t a g t t a g a t t t c a t a t a t a c t c a t t t g a a c c a g a c t g t

11,460 285 280 275 270 *WHKILSAGIEAIQRNREDM AmpR *WHKILSAGIEAIQRNREDM g t t a c c a a t g c t t a a t c a g t g a g g c a c c t a t c t c a g c g a t c t g t c t a t t t c g t t c a t c c a c a a t g g t t a c g a a t t a g t c a c t c c g t g g a t a g a g t c g c t a g a c a g a t a a a g c a a g t a g g t

11,520 265 260 255 250 TAQSGTTYIVVIRSPKGDPG AmpR TAQSGTTYIVVIRSPKGDPG t a g t t g c c t g a c t c c c c g t c g t g t a g a t a a c t a c g a t a c g g g a g g g c t t a c c a t c t g g c c a t c a a c g g a c t g a g g g g c a g c a c a t c t a t t g a t g c t a t g c c c t c c c g a a t g g t a g a c c g g

11,580 245 240 235 230 LAAIIGRSGREGAGSKDAIF AmpR LAAIIGRSGREGAGSKDAIF c c a g t g c t g c a a t g a t a c c g c g a g a c c c a c g c t c a c c g g c t c c a g a t t t a t c a g c a a t a a g g t c a c g a c g t t a c t a t g g c g c t c t g g g t g c g a g t g g c c g a g g t c t a a a t a g t c g t t a t t

11,640 225 220 215 210 WGAPLASRLLPGAVKDAEMW AmpR WGAPLASRLLPGAVKDAEMW a c c a g c c a g c c g g a a g g g c c g a g c g c a g a a g t g g t c c t g c a a c t t t a t c c g c c t c c a t c c t g g t c g g t c g g c c t t c c c g g c t c g c g t c t t c a c c a g g a c g t t g a a a t a g g c g g a g g t a g g

FspI 11,700 205 200 195 190 DILQQRSALTLLEGTLLKRL AmpR DILQQRSALTLLEGTLLKRL a g t c t a t t a a t t g t t g c c g g g a a g c t a g a g t a a g t a g t t c g c c a g t t a a t a g t t t g c g c a t c a g a t a a t t a a c a a c g g c c c t t c g a t c t c a t t c a t c a a g c g g t c a a t t a t c a a a c g c g t

11,760 185 180 175 170 TTAMAVPMTTDREDNPIAEN AmpR TTAMAVPMTTDREDNPIAEN a c g t t g t t g c c a t t g c t a c a g g c a t c g t g g t g t c a c g c t c g t c g t t t g g t a t g g c t t c a t t g c a a c a a c g g t a a c g a t g t c c g t a g c a c c a c a g t g c g a g c a g c a a a c c a t a c c g a a g t a

11,820 165 160 155 150 LEPEWRDLRTVHDGMNHLFA AmpR LEPEWRDLRTVHDGMNHLFA t c a g c t c c g g t t c c c a a c g a t c a a g g c g a g t t a c a t g a t c c c c c a t g t t g t g c a a a a a a g a g t c g a g g c c a a g g g t t g c t a g t t c c g c t c a a t g t a c t a g g g g g t a c a a c a c g t t t t t t c

PvuI 11,880 145 140 135 130 TLEKPGGITTLLLNAATNDS AmpR TLEKPGGITTLLLNAATNDS c g g t t a g c t c c t t c g g t c c t c c g a t c g t t g t c a g a a g t a a g t t g g c c g c a g t g t t a t c a c g c c a a t c g a g g a a g c c a g g a g g c t a g c a a c a g t c t t c a t t c a a c c g g c g t c a c a a t a g t g

11,940 125 120 115 110 MTIAASCLERVTMGDTLHKE AmpR MTIAASCLERVTMGDTLHKE t c a t g g t t a t g g c a g c a c t g c a t a a t t c t c t t a c t g t c a t g c c a t c c g t a a g a t g c t t t t a g t a c c a a t a c c g t c g t g a c g t a t t a a g a g a a t g a c a g t a c g g t a g g c a t t c t a c g a a a a

12,000 105 100 95 90 TVPSYEVLDNQSYHIRRGLQ AmpR TVPSYEVLDNQSYHIRRGLQ c t g t g a c t g g t g a g t a c t c a a c c a a g t c a t t c t g a g a a t a g t g t a t g c g g c g a c c g a g t t g a c a c t g a c c a c t c a t g a g t t g g t t c a g t a a g a c t c t t a t c a c a t a c g c c g c t g g c t c a a

12,060 85 80 75 70 EQGADIRSLVAGCLLVKFTS AmpR A T A A T A C C G C G C C A C A T A G C Amp-R EQGADIRSLVAGCLLVKFTS g c t c t t g c c c g g c g t c a a t a c g g g a t a a t a c c g c g c c a c a t a g c a g a a c t t t a a a a g t g c c g a g a a c g g g c c g c a g t t a t g c c c t a t t a t g g c g c g g t g t a t c g t c t t g a a a t t t t c a c g

12,120 65 60 55 50 MMPFREEPRFSELIKGSNLD AmpR MMPFREEPRFSELIKGSNLD t c a t c a t t g g a a a a c g t t c t t c g g g g c g a a a a c t c t c a a g g a t c t t a c c g c t g t t g a g a t a g t a g t a a c c t t t t g c a a g a a g c c c c g c t t t t g a g a g t t c c t a g a a t g g c g a c a a c t c t a

12,180 45 40 35 30 LEIYGVRAGLQDEADKVKVL AmpR LEIYGVRAGLQDEADKVKVL c c a g t t c g a t g t a a c c c a c t c g t g c a c c c a a c t g a t c t t c a g c a t c t t t t a c t t t c a c c a g g t c a a g c t a c a t t g g g t g a g c a c g t g g g t t g a c t a g a a g t c g t a g a a a a t g a a a g t g g t

12,240 25 TEPH 20 15 10 signal sequence AFVPLCFAAFFPILAV AmpR TEPHAFVPLCFAAFFPILAV g c g t t t c t g g g t g a g c a a a a a c a g g a a g g c a a a a t g c c g c a a a a a a g g g a a t a a g g g c g a c g c a a a g a c c c a c t c g t t t t t g t c c t t c c g t t t t a c g g c g t t t t t t c c c t t a t t c c c g c t

SspI 12,300 5 1 signal sequence RFHQISM AmpR AmpR promoter RFHQISM c a c g g a a a t g t t g a a t a c t c a t a c t c t t c c t t t t t c a a t a t t a t t g a a g c a t t t a t c a g g g t g c c t t t a c a a c t t a t g a g t a t g a g a a g g a a a a a g t t a t a a t a a c t t c g t a a a t a g t c c

12,360 AmpR promoter g t t a t t g t c t c a t g a g c g g a t a c a t a t t t g a a t g t a t t t a g a a a a a t a a a c a a a t a g g g g c a a t a a c a g a g t a c t c g c c t a t g t a t a a a c t t a c a t a a a t c t t t t t a t t t g t t t a t c c c c

SgrDI 12,420 AmpR promoter t t c c g c g c a c a t t t c c c c g a a a a g t g c c a c c t g a c g t c g a c g g a t c g g g a g a t c t c c c g a a a g g c g c g t g t a a a g g g g c t t t t c a c g g t g g a c t g c a g c t g c c t a g c c c t c t a g a g g g c t

12,480 A T G T T A G A C G A G A C T A C G G C pRS-marker t c c c c t a t g g t g c a c t c t c a g t a c a a t c t g c t c t g a t g c c g c a t a g t t a a g c c a g t a t c t a g g g g a t a c c a c g t g a g a g t c a t g t t a g a c g a g a c t a c g g c g t a t c a a t t c g g t c a t a g a

12,540 g c t c c c t g c t t g t g t g t t g g a g g t c g c t g a g t a g t g c g c g a g c a a a a t t t a a g c t a c a a c c g a g g g a c g a a c a c a c a a c c t c c a g c g a c t c a t c a c g c g c t c g t t t t a a a t t c g a t g t t g

12,600 a a g g c a a g g c t t g a c c g a c a a t t g c a t g a a g a a t c t g c t t a g g g t t a g g c g t t t t g c g c t t t c c g t t c c g a a c t g g c t g t t a a c g t a c t t c t t a g a c g a a t c c c a a t c c g c a a a a c g c g a

SpeI 12,660 CMV enhancer g c t t c g c g a t g t a c g g g c c a g a t a t a c g c g t t g a c a t t g a t t a t t g a c t a g t t a t t a a t a c g a a g c g c t a c a t g c c c g g t c t a t a t g c g c a a c t g t a a c t a a t a a c t g a t c a a t a a t t a t

12,720 CMV enhancer g t a a t c a a t t a c g g g g t c a t t a g t t c a t a g c c c a t a t a t g g a g t t c c g c g t t a c a t a a c t c a t t a g t t a a t g c c c c a g t a a t c a a g t a t c g g g t a t a t a c c t c a a g g c g c a a t g t a t t g a

12,780 CMV enhancer t a c g g t a a a t g g c c c g c c t g g c t g a c c g c c c a a c g a c c c c c g c c c a t t g a c g t c a a t a a t a t g c c a t t t a c c g g g c g g a c c g a c t g g c g g g t t g c t g g g g g c g g g t a a c t g c a g t t a t t a

12,840 CMV enhancer g a c g t a t g t t c c c a t a g t a a c g c c a a t a g g g a c t t t c c a t t g a c g t c a a t g g g t g g a g t a c t g c a t a c a a g g g t a t c a t t g c g g t t a t c c c t g a a a g g t a a c t g c a g t t a c c c a c c t c a t

12,900 CMV enhancer t t t a c g g t a a a c t g c c c a c t t g g c a g t a c a t c a a g t g t a t c a t a t g c c a a g t a c g c c c c c a a a t g c c a t t t g a c g g g t g a a c c g t c a t g t a g t t c a c a t a g t a t a c g g t t c a t g c g g g g g

12,960 CMV enhancer t a t t g a c g t c a a t g a c g g t a a a t g g c c c g c c t g g c a t t a t g c c c a g t a c a t g a c c t t a t g a t a a c t g c a g t t a c t g c c a t t t a c c g g g c g g a c c g t a a t a c g g g t c a t g t a c t g g a a t a c

SnaBI 3ʹ 5ʹ 13,013 CMV enhancer g g a c t t t c c t a c t t g g c a g t a c a t c t a c g t a t t a g t c a t c g c t a t t a c c a t g g c c t g a a a g g a t g a a c c g t c a t g t a g a t g c a t a a t c a g t a g c g a t a a t g g t a c c

Restriction Enzymes

Instructions: By default, all cutters are shown. Filter on number of cut sites or search by enzyme name.

Filter

Features

Primers

BLAST

BLAST (Basic Local Alignment Search Tool) finds regions of similarity between biological sequences. Click on the buttons below to submit a BLAST search to NCBI. The results will appear in a new window. See your recent BLAST results on NCBI's website.

  • Nucleotide-Nucleotide BLAST (BLASTN)

  • Translated Nucleotide-Protein BLAST (BLASTX)

  • Sequence alignment using BLAST (BLAST2)

Sequence Analyzer Guide

Map

Displays a graphical map based on nucleotide sequence data labeled with restriction enzymes, plasmid features, ORFs (theoretical open reading frames) and primers. Hovering over data labels will display additional information (e.g. cut site)

To select a portion of sequence, click one location on the plasmid and then a second location to display the sequence between the two locations.

Sequence

Displays both strands of base paired nucleotide sequences with annotated enzymes, plasmid features, ORFs (theoretical open reading frames) and primers. Hovering over data labels will display additional information (e.g. cut site).

To select a portion of sequence, click one location on the sequence and then a second location to display the sequence between the two locations.

Enzymes

List of restriction enzymes that can cut a given nucleotide sequence. Table lists enzyme name and the sequence location of the cut.

Features

List of common features detected in a given nucleotide sequence. Table lists feature name, location, size, color used to indicate its position on the map, and direction (if relevant).

Primers

List of commonly used primers detected in a given nucleotide sequence. Table lists primer name, sequence, length, binding site location, and direction.

BLAST

Use Basic Local Alignment Search Tool (BLAST) via the NCBI website to determine similarity between a given sequence and nucleotide (BLASTN) or protein (BLASTX) sequence databases. Additionally, align a custom nucleotide sequence against a given sequence using BLAST2.

File Downloads

GenBank

File contains the nucleotide sequence and annotated features in GenBank flat file format. Open the file with a text editor or plasmid mapping software to view the sequence.

SnapGene

File contains the nucleotide sequence and enhanced annotations from SnapGene Server. Open the file with SnapGene software or the free Viewer to view the plasmid map, sequence, and perform additional sequence analysis.