Quanfu Ma Lab: RBM24 interacts with RPL3 and regulates RPL3-eIF6 interaction

Unpublished

Plasmids from Article

ID	Plasmid	Purpose
208051	pcDNA3.1-Flag	Empty vector for mammalian expression, insert a Flag tag (with ATG) at XhoI site of pcDNA3.1 (-), xhoI site at 5' of flag tag destroyed, xhoI site at 3' of flag tag retained. XhoI in-frame with Flag
208052	pcDNA3.1-Flag-RBM24	Expressing N-ter Flag tagged human RBM24 (ENST00000379052) in mammalian cells
208053	pET28-3'TwStrep	Empty vector for prokaryotic expression, insert a Twin Strep tag (with TAA) between HindII and XhoI site of pET28a (+), both HindIII and XhoI are retained. HindIII in-frame with Twin Strep tag.
208616	pcDNA3.1-3'3xFlag	Empty vector for mammalian expression, insert 3xFlag tag using BamHI andAflII site of pcDNA3.1 (-), both site retained, BamHI in-frame with 3xFlag
208617	pcDNA3.1-StrepII	Empty vector for mammalian expression, insert a Strep II tag (with ATG) using XbaI and XhoI site of pcDNA3.1 (-), both site retained
208618	pcDNA3.1-3'TwStrep	Empty vector for mammalian expression, insert Twin Strep tag using BamHI and AflII site of pcDNA3.1 (-), both site retained, BamHI in-frame with Twin Strep tag
208619	pcDNA3.1-msEGFP1.8	Mammalian expression of a EGFP variant. Mutations were adopted from (Addgene#135301, PMC7508230), named msEGFP1.8 because N-ter and C-ter features of that in msGFP2 (PMC7508230) not included.
208620	pET28-5'TwStrep	Empty vector for prokaryotic expression, insert a Twin Strep tag between NdeI and BamHI site of pET28a (+), both site retained, T7 tag replaced, BamHI site in-frame with Twin Strep tag.
208621	pET28-Flag-RBM24	for E. coli expression of Flag tagged human RBM24 (ENST00000379052)
208622	pET28-mEGFP(A206K)-TwStrep	for E. coli expression of C-ter Twin Strep tagged mEGFP(A206K), EGFP cloned from pEGFP-C1, A206K introduced by seamless (Gibson) clone
208623	pET28-RPL3-Twstrep	for E. coli expression of C-ter Twin Strep tagged human RPL3 (ENST00000216146)
208624	pET28-RPL3-1~77-mEGFP(A206K)-TwStrep	for E. coli expression truncated human RPL3 (ENST00000216146), aa1~77, fused with mEGFP(A206K) and Twin Strep tag
208625	pET28-RPL3-72~197-mEGFP(A206K)-TwStrep	for E. coli expression of truncated human RPL3 (ENST00000216146), aa72~197, fused with mEGFP(A206K) and Twin Strep tag
208626	pET28-RPL3-191~307-mEGFP(A206K)-TwStrep	for E. coli expression of truncated human RPL3 (ENST00000216146), aa191~307, fused with mEGFP(A206K) and Twin Strep tag
208627	pET28-RPL3-299~398-mEGFP(A206K)-TwStrep	for E. coli expression of truncated human RPL3 (ENST00000216146), aa299~398, fused with mEGFP(A206K) and Twin Strep tag
208628	pET28-RPL3-1~41-mEGFP(A206K)-TwStrep	for E. coli expression of truncated human RPL3 (ENST00000216146), aa1~41, fused with mEGFP(A206K) and Twin Strep tag
208629	pET28-RPL3-212~285-mEGFP(A206K)-TwStrep	for E. coli expression of truncated human RPL3 (ENST00000216146), aa212~285, fused with mEGFP(A206K) and Twin Strep tag
208630	pET28-RPL3-Δ1~34,Δ215~284-TwStrep	for E. coli expression of truncated human RPL3 (ENST00000216146), aa35~398 with deletion of 215~284, fused with Twin Strep tag.
208632	pET28-RPL3-53~77-mEGFP(A206K)-TwStrep	for E. coli expression truncated human RPL3 (ENST00000216146), aa53~77, fused with mEGFP(A206K) and Twin Strep tag
208633	pET28-Flag-RBM24-1~93	for E. coli expression of truncated Flag tagged human RBM24 (ENST00000379052), aa1~93
208634	pET28-Flag-RBM24-95~236	for E. coli expression of truncated Flag tagged human RBM24 (ENST00000379052), aa95~236
208635	pET28-H12TEV	for E. coli expression of large protein, insert 6xhis + TEV site using NdeI and BamHI site in pET28a(+), note that it is 6xHis-Thrombin-6xHis-TEV, not continuous 12xHis. BamHI in-frame with tag.
208636	pET28-H12TEV-hEIF5B-1~1220(FL)	for E. coli expression of human eIF5B (ENST00000289371)
208637	pET28-H12TEV-hEIF5B-587~1220	for E. coli expression truncated human eIF5B (ENST00000289371), aa587~1220
208638	pET28-eIF4E-TwStrep	for E. coli expression of C-ter Twin Strep tagged human eIF4E (ENST00000450253)
208639	pUC-T7-J1~398-Luc	for in vitro transcription template preparation, insert T7 promoter, JFH1(AB047639)_1~398 and F-luc from pGL3-Basic, into pUC19
208640	pUC-T7-hTP53-P0~670-FLuc	for in vitro transcription template preparation, insert T7 promoter, TP53 translation initiation site P0 to +670 and F-luc from pGL3-Basic, into pUC19. P0~670 is confusing, BLAST is recommended.
208642	pUC-ODN	Empty vector for long single strand DNA (up to 1.5k) preparation. This plasmid is a pUC19 with 6 point mutations that remove BspQI, BtsI, BsrDI sites. Mutations adopted from Patent WO/2018/015995.
208643	pUC-ODN2	Empty vector for long single strand DNA (up to 1.5k) preparation. Further remove BsaI and BsmBI site in pUC-ODN (Addgene #208642).
208644	pUC-ODN-hsRPL3-CterTwStrep	for long single strand DNA donnor preparation for Twin Strep tag knock-in at C ter of human RPL3. Digest with nt.BspQI and nb.BsrDI to get ssDNA. This plasmid will cause re-edit and fail knock-in.
208645	pUC-ODN-hsRPL3-CterTwStrep-ΔPAM62&56	Remove PAM region in HomoArms by introduce SNP rs7495557 and rs760194863, to prevent cleavage after success edit. Digest with nt.BspQI and nb.BsrDI to get ssDNA.
208646	pcDNA3.1-CRISPR-Puro	pcDNA3.1 based empty vector for transient transfection of sgRNA-cas9. U6 promoter ~ puro Res cloned from lentiCRISPRv2 (Addgene#52961), lenti virus sequences not included.
208647	pcDNA3.1-CRISPR-Puro-sgRPL3-62	pcDNA3.1 based plasmid for transient transfection of sgRNA-cas9. Containing sgRNA targeting human RPL3 (GRCh38.p14_Chr22:39312968-39312987), 62 is a number given by www.e-crisp.org.
208649	pET28-RPL3L-TwStrep	for E. coli expression of C-ter Twin Strep tagged human RPL3L (ENST00000268661), contain SNP rs753338989, rs746044959, rs752134949
208979	pcDNA3.1-CRISPR-Puro-sgRPL3-56	pcDNA3.1 based plasmid for transient transfection of sgRNA-cas9. Containing sgRNA targeting human RPL3 (GRCh38.p14_Chr22:39312868-39312887), 56 is a number given by www.e-crisp.org.