LOCUS       sequence-238578-        6307 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression plasmid for wild-type VCre with an N-terminal 
            His-tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6307)
  AUTHORS   Bessen JL, Afeyan LK, Dancik V, Koblan LW, Thompson DB, Leichner C, 
            Clemons PA, Liu DR
  TITLE     High-resolution specificity profiling and off-target prediction for 
            site-specific DNA recombinases.
  JOURNAL   Nat Commun. 2019 Apr 26;10(1):1937. doi: 10.1038/s41467-019-09987-0.
  PUBMED    31028261
REFERENCE   2  (bases 1 to 6307)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2019-04-26"; volume: "10(1):1937. doi"; pages: " 
            10.1038/s41467-019-09987-0"
COMMENT     Sequence Label: pET-His-VCre
FEATURES             Location/Qualifiers
     source          1..6307
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     76..95
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     183..202
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        183..201
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    202..226
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /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             241..263
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             276..293
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1455..1499
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(1537..1555)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1551..1598
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1635..2090
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1722..1741)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1932..1953
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(join(2205..2996,2997..3065))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     2828..2847
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3066..3158)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3239..3827
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3728..3747
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3981..3998
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4013..4152
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4078..4097)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4238..4260
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4254..4445)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5017..6099)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=lacI
                     /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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     6061..6080
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6100..6177)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 cacggggcct gccaccatac ccacgccgaa acaagcgctc atgagcccga agtggcgagc
       61 ccgatcttcc acatcggtga tgtcggcgat ataggcgcca gcaaccgcac ctgtggcgcc
      121 ggtgatgccg gccacgatgc gtccggcgta gaggatcgag atcgatctcg atcccgcgaa
      181 attaatacga ctcactatag gggaattgtg agcggataac aattcccctc tagaaataat
      241 tttgtttaac tttaagaagg agatatacca tggcgcatca ccaccaccat cacggcggga
      301 gcatagagaa tcagctaagc ttactgggcg atttttctgg agtacgccct gacgatgtaa
      361 aaacggctat ccaggctgcg cagaaaaaag gaattaatgt cgccgaaaat gaacagttta
      421 aagccgcgtt tgagcacctg ctgaatgagt tcaagaaacg cgaggaacgt tatagtccaa
      481 atactctcag gcggttagag tcggcgtgga cttgctttgt ggattggtgt cttgccaacc
      541 atcgacattc tctgccggca actccagata cagtagaagc gttcttcatt gagagagcag
      601 aggagcttca tcgtaacacc ttatcagttt atcgttgggc aatttcgcgg gtgcaccgcg
      661 tagccggttg ccctgatccg tgtctagata tatatgtaga agatcgtcta aaggctatag
      721 ctcggaagaa agtgcgcgaa ggtgaagctg tcaaacaggc tagtcccttt aacgagcagc
      781 accttctgaa gctgactagt ctgtggtatc gaagtgataa actcctgctt aggcgcaatt
      841 tagcattgct ggccgtggct tacgagtcta tgttacgggc atcggaactg gcgaacatcc
      901 gagtgtccga tatggaactt gctggagatg gtacagcgat ccttaccatt cctattacga
      961 aaacaaatca ctccggtgaa cccgacacat gtattctctc acaggacgtt gtctctctgc
     1021 tgatggacta tacggaggca ggcaaactcg atatgtctag cgacggtttt ctatttgttg
     1081 gggtcagtaa acataatacg tgtatcaaac cgaagaagga taaacaaaca ggcgaagtgc
     1141 ttcacaaacc tattacgaca aagacggtag aaggagtctt ctattcggca tgggaaacct
     1201 tagacttggg aaggcaaggg gtgaaacctt ttactgcaca ctctgcccgt gtgggagccg
     1261 cacaggatct gctgaagaag ggctataaca ccttacagat ccagcagagc gggagatgga
     1321 gctctggcgc aatggtggcc cgttacggtc gtgctattct ggcccgagat ggcgccatgg
     1381 cacattctag agtaaaaaca cgttccgcgc caatgcaatg gggcaaagat gaaaaggact
     1441 aactcgagtc tggtaaagaa accgctgctg cgaaatttga acgccagcac atggactcgt
     1501 ctactagcgc agcttaatta acctaggctg ctgccaccgc tgagcaataa ctagcataac
     1561 cccttggggc ctctaaacgg gtcttgaggg gttttttgct gaaaggagga actatatccg
     1621 gattggcgaa tgggacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     1681 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     1741 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
     1801 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
     1861 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
     1921 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta
     1981 ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat
     2041 ttaacaaaaa tttaacgcga attttaacaa aatattaacg tttacaattt ctggcggcac
     2101 gatggcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     2161 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     2221 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     2281 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     2341 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     2401 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     2461 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     2521 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     2581 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     2641 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     2701 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     2761 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     2821 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     2881 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     2941 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     3001 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     3061 ctcatactct tcctttttca atcatgattg aagcatttat cagggttatt gtctcatgag
     3121 cggatacata tttgaatgta tttagaaaaa taaacaaata ggtcatgacc aaaatccctt
     3181 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     3241 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     3301 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     3361 gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc caccacttca
     3421 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     3481 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     3541 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     3601 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     3661 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     3721 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     3781 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     3841 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
     3901 tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc
     3961 gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcctgatgc
     4021 ggtattttct ccttacgcat ctgtgcggta tttcacaccg caatggtgca ctctcagtac
     4081 aatctgctct gatgccgcat agttaagcca gtatacactc agctatcgct acgtgactgg
     4141 gtcatggctg cgccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg
     4201 ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg
     4261 ttttcaccgt catcaccgaa acgcgcgagg cagctgcggt aaagctcatc agcgtggtcg
     4321 tgaagcgatt cacagatgtc tgcctgttca tccgcgtcca gctcgttgag tttctccaga
     4381 agcgttaatg tctggcttct gataaagcgg gccatgttaa gggcggtttt ttcctgtttg
     4441 gtcactgatg cctccgtgta agggggattt ctgttcatgg gggtaatgat accgatgaaa
     4501 cgagagagga tgctcacgat acgggttact gatgatgaac atgcccggtt actggaacgt
     4561 tgtgagggta aacaactggc ggtatggatg cggcgggacc agagaaaaat cactcagggt
     4621 caatgccagc gcttcgttaa tacagatgta ggtgttccac agggtagcca gcagcatcct
     4681 gcgatgcaga tccggaacat aatggtgcag ggcgctgact tccgcgtttc cagactttac
     4741 gaaacacgga aaccgaagac cattcatgtt gttgctcagg tcgcagacgt tttgcagcag
     4801 cagtcgcttc acgttcgctc gcgtatcggt gattcattct gctaaccagt aaggcaaccc
     4861 cgccagccta gccgggtcct caacgacagg agcacgatca tgctagtcat gccccgcgcc
     4921 caccggaagg agctgactgg gttgaaggct ctcaagggca tcggtcgaga tcccggtgcc
     4981 taatgagtga gctaacttac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     5041 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     5101 attgggcgcc agggtggttt ttcttttcac cagtgagacg ggcaacagct gattgccctt
     5161 caccgcctgg ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc ccagcaggcg
     5221 aaaatcctgt ttgatggtgg ttaacggcgg gatataacat gagctgtctt cggtatcgtc
     5281 gtatccaact accgagatgt ccgcaccaac gcgcagcccg gactcggtaa tggcgcgcat
     5341 tgcgcccagc gccatctgat cgttggcaac cagcatcgca gtgggaacga tgccctcatt
     5401 cagcatttgc atggtttgtt gaaaaccgga catggcactc cagtcgcctt cccgttccgc
     5461 tatcggctga atttgattgc gagtgagata tttatgccag ccagccagac gcagacgcgc
     5521 cgagacagaa cttaatgggc ccgctaacag cgcgatttgc tggtgaccca atgcgaccag
     5581 atgctccacg cccagtcgcg taccgtcttc atgggagaaa ataatactgt tgatgggtgt
     5641 ctggtcagag acatcaagaa ataacgccgg aacattagtg caggcagctt ccacagcaat
     5701 ggcatcctgg tcatccagcg gatagttaat gatcagccca ctgacgcgtt gcgcgagaag
     5761 attgtgcacc gccgctttac aggcttcgac gccgcttcgt tctaccatcg acaccaccac
     5821 gctggcaccc agttgatcgg cgcgagattt aatcgccgcg acaatttgcg acggcgcgtg
     5881 cagggccaga ctggaggtgg caacgccaat cagcaacgac tgtttgcccg ccagttgttg
     5941 tgccacgcgg ttgggaatgt aattcagctc cgccatcgcc gcttccactt tttcccgcgt
     6001 tttcgcagaa acgtggctgg cctggttcac cacgcgggaa acggtctgat aagagacacc
     6061 ggcatactct gcgacatcgt ataacgttac tggtttcaca ttcaccaccc tgaattgact
     6121 ctcttccggg cgctatcatg ccataccgcg aaaggttttg cgccattcga tggtgtccgg
     6181 gatctcgacg ctctccctta tgcgactcct gcattaggaa gcagcccagt agtaggttga
     6241 ggccgttgag caccgccgcc gcaaggaatg gtgcatgcaa ggagatggcg cccaacagtc
     6301 ccccggc
//
