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Species: Synthetic
Genetic Insert: Nsp3e_NAB
Vector Backbone Description: Backbone Size:6017; Vector Backbone:pETM33; Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:37704626
Comments: insert amino acid sequence: GKKDNSYFTEQPIDLVPNQPYPNASFDNFKFVCDNIKFADDLNQLTGYKKPASRELKVTFFPDLNGDVVAIDYKHYTPSFKKGAKLLHKPIVWHVNNATNKATYKPNTWCIRCLWSTKPVETx
Proper citation: RRID:Addgene_156473 Copy
Species: Synthetic
Genetic Insert: Nsp15
Vector Backbone Description: Backbone Size:6017; Vector Backbone:pETM33; Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:37704626
Comments: insert amino acid sequence: GLQSLENVAFNVVNKGHFDGQQGEVPVSIINNTVYTKVDGVDVELFENKTTLPVNVAFELWAKRNIKPVPEVKILNNLGVDIAANTVIWDYKRDAPAHISTIGVCSMTDIAKKPTETICAPLTVFFDGRVDGQVDLFRNARNGVLITEGSVKGLQPSVGPKQASLNGVTLIGEAVKTQFNYYKKVDGVVQQLPETYFTQSRNLQEFKPRSQMEIDFLELAMDEFIERYKLEGYAFEHIVYGDFSHSQLGGLHLLIGLAKRFKESPFELEDFIPMDSTVKNYFITDAQTGSSKCVCSVIDLLLDDFVEIIKSQDLSVVSKVVKVTIDYTEISFMLWCKDGHVETFYPKLx
Proper citation: RRID:Addgene_156481 Copy
Species: Homo sapiens
Genetic Insert: Fibroblast growth factor 2
Vector Backbone Description: Backbone Size:5369; Vector Backbone:pHis; Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:32383752
Proper citation: RRID:Addgene_157657 Copy
Species: Homo sapiens
Genetic Insert: Apoptosis inhibitor 5
Vector Backbone Description: Backbone Size:5369; Vector Backbone:pET28b; Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:32383752
Proper citation: RRID:Addgene_157655 Copy
Species: Other
Genetic Insert: Cohesin (R.f) with ddFLN4, ELP, Histag and ybbR tag
Vector Backbone Description: Backbone Size:5200; Vector Backbone:pET28a; Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:33191756
Proper citation: RRID:Addgene_157672 Copy
Species: Batis maritima
Genetic Insert: MHT
Vector Backbone Description: Vector Backbone:pET28b; Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:33104325
Proper citation: RRID:Addgene_157639 Copy
Genetic Insert: sMHT(1-113) -CheA
Vector Backbone Description: Vector Backbone:pET28b; Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:33104325
Proper citation: RRID:Addgene_157642 Copy
Genetic Insert: sMHT(1-113)
Vector Backbone Description: Vector Backbone:pET28b; Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:33104325
Proper citation: RRID:Addgene_157641 Copy
Genetic Insert: sMHT(1-113)-SYNZIP17
Vector Backbone Description: Vector Backbone:pET28b; Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:33104325
Proper citation: RRID:Addgene_157640 Copy
Vector Backbone Description: Backbone Marker:Clontech; Backbone Size:3939; Vector Backbone:pEGFP-C1; Vector Types:Mammalian Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:31317736
Proper citation: RRID:Addgene_157777 Copy
Vector Backbone Description: Backbone Size:5401; Vector Backbone:pelB-MBP (DNASU access. no. EvNO00813783); Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Comments: - Empty vector (kanamycin-R) adds TEV-protease-removable N-terminal PelB signal sequence + His10, plus potential C-term His6. Non-leaky expression due to T7lac promoter.
- Has an advantage in leaving no added amino acid residues following TEV protease cleavage of PelB signal sequence + His10. This advantage is made possible by cloning into the two BseRI restriction sites.
- Subcloning requires ligation-independent cloning and use of the two BseRI restriction sites that are present in this empty vector. The two BseRI sites span a removable 432 nucleotide sequence. Cloning into the BseRI-digested vector can be achieved using a ligation-independent system that is dependent on homologous recombination, such as the Takara In-Fusion HD Cloning Plus system, which requires simply designing the PCR-derived insert (containing the protein of interest) to contain 15 bp at each end that matches the 15 bp at each end of the linearized parent vector. In other words, add these 15 bp, AACCTGTACTTCCAA, to the 5' end of the PCR FORWARD primer, and add these 15 bp, GTGGTGGTGCTCGAG, to the 5' end of the PCR REVERSE primer.
- The sequence of the added N-terminus is the PelB signal sequence (M1-A22 of GenBank #AAA24848) + 13 aa linker + His10-tag + 4 aa linker + TEV protease cleavage site. The N-terminal protein sequence is: MKYLLPTAAAGLLLLAAQPAMA + MDIGINSDPNSSS + HHHHHHHHHH + PMGS + ENLYFQ*X, where * is the TEV protease cleavage site and X is the N-terminal residue of the introduced protein of interest. Note that TEV protease cleaves most efficiently when X = S, G, A, M; for other compatible residues see Kapust et al 2002, PMID 12074568.
- A C-terminal His6-tag is possible: if no stop codon is included in the cloned insert, the sequence LEHHHHHH is added to the C-terminus.
- The PelB signal sequence directs the protein of interest to the E. coli inner membrane. Whether the protein traverses the inner membrane is dependent on the protein of interest.
- BseRI cuts 8-10 nucleotides outside of its own recognition sequence and is a non-palindromic site. The placement & orientation of the two BseRI sites eliminates all of the BseRI recognition sequence in the resulting subclone.
- To verify the PCR-generated insert sequence, use sequencing primers T7 (TAATACGACTCACTATAGGG), T7-Ter (GCTAGTTATTGCTCAGCGG).
Proper citation: RRID:Addgene_157741 Copy
Species: Homo sapiens
Genetic Insert: AURKA
Vector Backbone Description: Backbone Marker:Clontech; Backbone Size:4733; Vector Backbone:pEGFP-N1; Vector Types:Mammalian Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:30070631
Proper citation: RRID:Addgene_157755 Copy
Species: SARS-CoV-2
Genetic Insert: N
Vector Backbone Description: Vector Backbone:pTHMT; Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:33200826
Proper citation: RRID:Addgene_157869 Copy
Species: Homo sapiens
Genetic Insert: hnRNPA2
Vector Backbone Description: Vector Backbone:pJ411; Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:32870271
Proper citation: RRID:Addgene_157866 Copy
Species: Homo sapiens
Genetic Insert: hnRNPA2
Vector Backbone Description: Vector Backbone:pJ411; Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:32870271
Proper citation: RRID:Addgene_157865 Copy
Species: SARS-CoV-2
Genetic Insert: N
Vector Backbone Description: Vector Backbone:pTHMT; Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:33200826
Comments: Please note: This plasmid contains a D15H mutation in MBP. This mutation is not known to affect plasmid function.
Proper citation: RRID:Addgene_157872 Copy
Species: SARS-CoV-2
Genetic Insert: N
Vector Backbone Description: Vector Backbone:pTHMT; Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:33200826
Proper citation: RRID:Addgene_157871 Copy
Species: Synthetic
Genetic Insert: de novo designed protein RO2_10
Vector Backbone Description: Vector Backbone:pET-28a(+); Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:32855341
Comments: Please visit https://www.biorxiv.org/content/10.1101/2020.04.14.041772v1 for bioRxiv preprint.
Proper citation: RRID:Addgene_158019 Copy
Species: Synthetic
Genetic Insert: de novo designed protein RO2_9
Vector Backbone Description: Vector Backbone:pET-28a(+); Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:32855341
Comments: Please visit https://www.biorxiv.org/content/10.1101/2020.04.14.041772v1 for bioRxiv preprint.
Proper citation: RRID:Addgene_158018 Copy
Species: Synthetic
Genetic Insert: de novo designed protein RO1_8
Vector Backbone Description: Vector Backbone:pET-28a(+); Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:32855341
Comments: Please visit https://www.biorxiv.org/content/10.1101/2020.04.14.041772v1 for bioRxiv preprint.
Proper citation: RRID:Addgene_158013 Copy
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