...repeatedly the reactive groups on a nucleotide when adding a nucleotide one at a time. The main property ...more than one nucleotide at that position termed a mixed site. One can also use a nucleotide-based molecular... Gel Electrophoresis Primer Design for PCR Oligonucleotide primers are necessary when running a PCR reaction.... They are synthesized chemically by joining nucleotides together. One must selectively block and unblock... the primer usually has stretches of several nucleotides. Also, both of the 3’ ends of the hybridized ...process. When designing, if unsure about what nucleotide to put at a certain position within the primer...molecular insert (inosine) instead of a regular nucleotide for broader pairing capabilities. Taking into ...

...will include the free nucleotide dGTP in the reaction (exclude all other nucleotides from standard polymerase...without ligation. Because of its dual polymerase/exonuclease functions, T4 DNA polymerase can create overhangs... which a single dNTP is included to stop the exonuclease function of the enzyme and shift its activity...polymerase protocol), causing the enzyme to perform exonuclease activity until the first "G" in the sequence.... the polymerase reaction is favored over the exonuclease reaction, the polymerase will add back the guanosine...manufacturer. It is very important to remove all free nucleotides from your PCR product before proceeding, as they... they will interfere with the exonuclease activity of T4 Pol in the following step. This may be accomplished...

...DNA and the Taq polymerase, PCR requires free nucleotides [dNTPs; adenine (A), cytosine (C), guanine (G...also requires two unique single stranded DNA oligonucleotide (oligo) primers, which anneal to the regions...and the hydrogen bonds are broken between the nucleotide base pairs. The initial denaturation step is ...increases the error rate of the DNA polymerase. Deoxynucleoside Triphosphates (dNTPs): Are the building blocks...strands of the template DNA. They are strands of nucleic acid that are starting points for DNA elongation...

...primer should be a minimum of 50 nucleotides and a maximum of 300 nucleotides from your target. Are there ...etc.) involves selecting one or more unique oligonucleotide primers that flank the regions of your plasmid...colored peaks corresponding to one of the four nucleotide bases. This is an example of a trace file from...

...ssDNA ends from the ssDNA-specific endonuclease activity of T5 Exonuclease and furthermore reduces secondary...specific and unique function for the reaction: T5 Exonuclease - creates single-strand DNA 3’ overhangs by chewing... High-Fidelity DNA Polymerase - incorporates nucleotides to “fill in” the gaps in the annealed DNA fragments...

...standard curve RNase-free DNase 10X DNase buffer Nuclease-free water Microcentrifuge tubes 96-well optical...0.15 μL 15 μL 100 μM Reverse Primer 0.15 μL 15 μL Nuclease Free Water 4.7 μL 470 μL Procedure Prepare a plasmid...10 9 stock or previous dilution (μL) Volume of nuclease-free water (uL) Molecules per μL 10 90 2 x 10 ...Dilution Series Volume of sample (uL) Volume of nuclease free water (uL) Dilution factor Total dilution...

... 5 minutes to allow for dissociation of the nucleoprotein complexes. The effectiveness of your RNA isolation...agarose gel, or bioanalyzer. For more information on nucleic acid quantification, see our protocol for DNA quantification...

...improves transduction efficiency by facilitating nuclear import of the vector’s preintegration complex in...found, search for NAR(N 17 )YNN, where N is any nucleotide, R is a purine (A,G), and Y is a pyrimidine (...targeting introns. Avoid stretches of 4 or more nucleotide repeats, especially repeated Ts because polyT...program. Select sequences that have at least 3 nucleotide mismatches to all unrelated genes. TIP: Addgene...

...your substance maximally absorbs. In the case of nucleic acid (DNA and RNA), the maximal absorbance is at...Protein maximally absorbs at 280nm and the ratio of nucleic acid to protein (260/280) is generally used as ...

... 37 °C for 5 min. Note: Ribonuclease A (RNase A) is a pancreatic ribonuclease that digests single-stranded...

...targets. Primers and a probe against the cellular ribonuclease P/MRP 30 kDa subunit (RPP30) are used as a control...primers/probe (HEX/VIC) 1 µL 9 µL 900 nM, 250 nM Nuclease-free water 4 µL 36 µL Total Volume 16 µL 144 µL...

...Ligation Dilute 5μL of annealed oligos with 45μL nuclease-free water and quantify the concentration (should...

...Restriction enzymes are naturally occurring bacterial endonucleases that recognize a large range of DNA sequences...

... the plasmid should also be propagated in an endonuclease negative E. coli strain such as NEB stable. ...

...phosphodiester linkages, which permanently join the nucleotides together. After ligation, the insert DNA is physically...

... on a rocking platform. (Optional) Counterstain nuclei with 500 µL of 300 nM DAPI working solution for...

...the plasmids should also be propagated in an endonuclease negative E. coli strain such as NEB stable. ...

...use a HEK293 line stable expressing Epstein Barr nuclear antigen (EBNA). Safety Warnings HEK293 cells are...

... hard to extract proteins such as those in the nucleus. Incubate on ice for 30 min . Centrifuge lysate...

...Reverse ITR Primer (10 µM) 1.8 µL 16.2 µL 900 nM Nuclease-free water 5.9 µL 71.55 µL Total Volume 20 µL ...