Basic Vector Information
pTYB21 vector Vector Map
Plasmid Resuspension Protocol:
1. Centrifuge at 5,000×g for 5 min.
2. Carefully open the tube and add 20 μl of sterile water to dissolve the DNA.
3. Close the tube and incubate for 10 minutes at room temperature.
4. Briefly vortex the tube and then do a quick spin to concentrate the liquid at the bottom. Speed is less than 5000×g.
5.Store the plasmid at -20 ℃.
pTYB21 vector Sequence
LOCUS pTYB21. 7514 bp DNA circular SYN 01-JAN-1980 DEFINITION Bacterial expression vector that encodes a thiol-cleavable N-terminal intein with a chitin-binding domain. ACCESSION . VERSION . KEYWORDS pTYB21. SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7514) AUTHORS Chong S, Montello GE, Zhang A, Cantor EJ, Liao W, Xu MQ, Benner J. TITLE Utilizing the C-terminal cleavage activity of a protein splicing element to purify recombinant proteins in a single chromatographic step. JOURNAL Nucleic Acids Res. 1998;26:5109-15. PUBMED 9801307 REFERENCE 2 (bases 1 to 7514) AUTHORS New England Biolabs TITLE Direct Submission REFERENCE 3 (bases 1 to 7514) AUTHORS . TITLE Direct Submission COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Nucleic Acids Res."; date: "1998"; volume: "26"; pages: "5109-15" COMMENT SGRef: number: 2; type: "Journal Article" COMMENT Clone with the SapI site to generate a fusion protein that can be isolated with no added amino acids. FEATURES Location/Qualifiers source 1..7514 /mol_type="other DNA" /organism="synthetic DNA construct" promoter 35..139 /label=AmpR promoter CDS 140..997 /label=AmpR /note="beta-lactamase" rep_origin complement(1042..1555) /direction=LEFT /label=M13 ori /note="M13 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" rep_origin 1666..2254 /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(2626..2814) /label=rop /note="Rop protein, which maintains plasmids at low copy number" protein_bind complement(3337..3358) /label=CAP binding site /note="CAP binding activates transcription in the presence of cAMP." CDS complement(3374..4453) /label=lacI /note="lac repressor" promoter complement(4454..4531) /label=lacI promoter terminator 4684..4770 /label=rrnB T1 terminator /note="transcription terminator T1 from the E. coli rrnB gene" terminator 4867..4953 /gene="Escherichia coli rrnB" /label=Escherichia coli rrnB terminator /note="rrnB T1 terminator" /note="transcription terminator T1 from the E. coli rrnB gene" terminator 5050..5136 /gene="Escherichia coli rrnB" /label=Escherichia coli rrnB terminator /note="rrnB T1 terminator" /note="transcription terminator T1 from the E. coli rrnB gene" terminator 5233..5319 /gene="Escherichia coli rrnB" /label=Escherichia coli rrnB terminator /note="rrnB T1 terminator" /note="transcription terminator T1 from the E. coli rrnB gene" terminator complement(5419..5462) /label=rrnB T1 terminator /note="transcription terminator T1 from the E. coli rrnB gene" promoter 5637..5655 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" protein_bind 5656..5680 /label=lac operator /note="The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-beta-D-thiogalactopyranoside (IPTG)." RBS 5695..5717 /label=RBS /note="efficient ribosome binding site from bacteriophage T7 gene 10 (Olins and Rangwala, 1989)" CDS 5725..5754 /codon_start=1 /product="N-terminal peptide from mature maltose-binding protein" /label=N-terminal peptide from mature maltose-binding /note="MBP peptide" /translation="MKIEEGKLVI" CDS 5770..6588 /label=Sce VMA intein 5' region /note="5' region of the intein from the yeast Vma1 subunit of the vacuolar ATPase (Chong et al., 1998)" CDS 6592..6747 /label=CBD /note="chitin binding domain from chitinase A1 (Watanabe et al., 1994)" CDS 6751..7299 /label=Sce VMA intein 3' region /note="modified 3' region of the intein from the yeast Vma1 subunit of the vacuolar ATPase (Chong et al., 1998)" misc_feature 7300..7377 /label=MCS /note="MCS" /note="multiple cloning site" terminator 7442..7489 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase"
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