Basic Vector Information
- Vector Name:
- pNIC-CTHF
- Antibiotic Resistance:
- Kanamycin
- Length:
- 7260 bp
- Type:
- Structural Genomics Vectors
- Replication origin:
- ori
- Source/Author:
- Savitsky P, Bray J, Cooper CD, Marsden BD, Mahajan P, Burgess-Brown
- Copy Number:
- High copy number
- Promoter:
- sacB
pNIC-CTHF 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 ℃.
pNIC-CTHF vector Sequence
LOCUS 40924_33227 7260 bp DNA circular SYN 18-DEC-2018 DEFINITION Expression vector pNIC-CTHF, complete sequence. ACCESSION . VERSION . KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7260) AUTHORS Gileadi O, Burgess-Brown N, Loppnau P. TITLE Vectors for Ligation-independent Cloning (LIC) JOURNAL Unpublished REFERENCE 2 (bases 1 to 7260) AUTHORS Gileadi O, Burgess-Brown N. TITLE Direct Submission JOURNAL Submitted (27-DEC-2006) Structural Genomics Consortium, University of Oxford, Botnar Research Centre, Oxford OX3 7LD, UK REFERENCE 3 (bases 1 to 7260) TITLE Direct Submission REFERENCE 4 (bases 1 to 7260) AUTHORS . TITLE Direct Submission COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Unpublished" COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted (27-DEC-2006) Structural Genomics Consortium, University of Oxford, Botnar Research Centre, Oxford OX3 7LD, UK" COMMENT SGRef: number: 3; type: "Journal Article" FEATURES Location/Qualifiers source 1..7260 /mol_type="other DNA" /organism="synthetic DNA construct" promoter 1..19 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" protein_bind 20..44 /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 59..81 /label=RBS /note="efficient ribosome binding site from bacteriophage T7 gene 10 (Olins and Rangwala, 1989)" promoter 96..528 /label=sacB promoter /note="sacB promoter and control region" CDS 529..1947 /codon_start=1 /label=SacB /note="secreted levansucrase that renders bacterial growth sensitive to sucrose" /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH ITRHDMLQIPEQQKNEKYKVPEFDSSTIKNISSAKGLDVWDSWPLQNTDGTVANYHGYH IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV VKDSILEQGQLTVNK" CDS 2009..2029 /codon_start=1 /label=TEV site /note="tobacco etch virus (TEV) protease recognition and cleavage site" /translation="ENLYFQS" CDS 2030..2047 /codon_start=1 /label=6xHis /note="6xHis affinity tag" /translation="HHHHHH" CDS 2048..2071 /codon_start=1 /label=FLAG /note="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /translation="DYKDDDDK" CDS 2121..2138 /codon_start=1 /label=6xHis /note="6xHis affinity tag" /translation="HHHHHH" terminator 2205..2252 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" rep_origin 2289..2744 /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" CDS complement(2840..3652) /codon_start=1 /label=KanR /note="aminoglycoside phosphotransferase" /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF" rep_origin 3774..4362 /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" misc_feature complement(4548..4690) /label=bom /note="basis of mobility region from pBR322" CDS complement(4795..4983) /codon_start=1 /label=rop /note="Rop protein, which maintains plasmids at low copy number" /translation="VTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA DELYRSCLARFGDDGENL" protein_bind complement(5758..5779) /label=CAP binding site /note="CAP binding activates transcription in the presence of cAMP." CDS complement(5795..6874) /codon_start=1 /label=lacI /note="lac repressor" /translation="VKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR ALADSLMQLARQVSRLESGQ" promoter complement(6875..6952) /label=lacI promoter
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