LOCUS       sequence-319837-        5992 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial Expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5992)
  TITLE     SGC Toronto 2019-Q1 to 2020-Q2
REFERENCE   2  (bases 1 to 5992)
  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"
COMMENT     Sequence Label: 6VAH
FEATURES             Location/Qualifiers
     source          1..5992
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             746..763
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             813..830
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(883..901)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      897..944
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      981..1436
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1068..1087)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1278..1299
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1528..2343)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1528..2343)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2251..2270
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2335..2354
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2465..3053
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2954..2973
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3207..3224
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3239..3378
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3304..3323)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3464..3486
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3480..3671)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4480..5562)
                     /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     5524..5543
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5563..5640)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5846..5865
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5949..5968
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5949..5967
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5968..5992
                     /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)."
ORIGIN
        1 cctctagaaa taattttgtt taaccttaag aaggagatat actatggcct ggcaggctcg
       61 gggcctgggc accgcccggt tgcagctggt agagttctca gccttcgtgg aaccgccaga
      121 tgcagttgat tcttaccaga ggcacctgtt cgtgcacatc agccagcact gccccagccc
      181 cggagcgccg ccgctcgaga gtgtggacgt ccggcagatc tacgacaaat tccctgagaa
      241 aaagggtggc ctccgagagc tatatgatcg tggccccccc catgccttct tcctggtcaa
      301 gttctgggcg gacctgaact ggggcccaag tggtgaggag gcaggggccg gtggcagcat
      361 cagcagtggt ggcttctacg gagtgagcag ccagtatgag agcctggaac acatgaccct
      421 cacctgttcc tccaaggtct gctcttttgg caagcaggtg gtggagaagg tggagacgga
      481 acgggcccag ctggaggacg gcagatttgt gtaccgcctg ctgcgctcgc ccatgtgcga
      541 gtacctggtg aatttcttgc acaagttgcg gcagctgcct gagcgataca tgatgaacag
      601 cgtcctggaa aacttcacca tcctccaggt ggtgacaaac agagacaccc aggaactgct
      661 gctctgcacc gcctatgtct tcgaggtctc caccagcgag cgtggggccc agcatcacat
      721 ttaccgcctg gtcagggacg tggagcacca tcatcaccac cattgaggat ccgaattcga
      781 gctccgtcga caagcttgcg gccgcactcg agcaccacca ccaccaccac tgagatccgg
      841 ctgctaacaa agcccgaaag gaagctgagt tggctgctgc caccgctgag caataactag
      901 cataacccct tggggcctct aaacgggtct tgaggggttt tttgctgaaa ggaggaacta
      961 tatccggatt ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
     1021 ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt
     1081 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
     1141 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg
     1201 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
     1261 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc
     1321 ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga
     1381 gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgctta caatttaggt
     1441 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     1501 aatatgtatc cgctcatgaa ttaattctta gaaaaactca tcgagcatca aatgaaactg
     1561 caatttattc atatcaggat tatcaatacc atatttttga aaaagccgtt tctgtaatga
     1621 aggagaaaac tcaccgaggc agttccatag gatggcaaga tcctggtatc ggtctgcgat
     1681 tccgactcgt ccaacatcaa tacaacctat taatttcccc tcgtcaaaaa taaggttatc
     1741 aagtgagaaa tcaccatgag tgacgactga atccggtgag aatggcaaaa gtttatgcat
     1801 ttctttccag acttgttcaa caggccagcc attacgctcg tcatcaaaat cactcgcatc
     1861 aaccaaaccg ttattcattc gtgattgcgc ctgagcgaga cgaaatacgc gatcgctgtt
     1921 aaaaggacaa ttacaaacag gaatcgaatg caaccggcgc aggaacactg ccagcgcatc
     1981 aacaatattt tcacctgaat caggatattc ttctaatacc tggaatgctg ttttcccggg
     2041 gatcgcagtg gtgagtaacc atgcatcatc aggagtacgg ataaaatgct tgatggtcgg
     2101 aagaggcata aattccgtca gccagtttag tctgaccatc tcatctgtaa catcattggc
     2161 aacgctacct ttgccatgtt tcagaaacaa ctctggcgca tcgggcttcc catacaatcg
     2221 atagattgtc gcacctgatt gcccgacatt atcgcgagcc catttatacc catataaatc
     2281 agcatccatg ttggaattta atcgcggcct agagcaagac gtttcccgtt gaatatggct
     2341 cataacaccc cttgtattac tgtttatgta agcagacagt tttattgttc atgaccaaaa
     2401 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     2461 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     2521 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     2581 gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc
     2641 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     2701 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     2761 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     2821 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     2881 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     2941 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     3001 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     3061 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     3121 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
     3181 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc
     3241 tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcaat ggtgcactct
     3301 cagtacaatc tgctctgatg ccgcatagtt aagccagtat acactccgct atcgctacgt
     3361 gactgggtca tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct
     3421 tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt
     3481 cagaggtttt caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag ctcatcagcg
     3541 tggtcgtgaa gcgattcaca gatgtctgcc tgttcatccg cgtccagctc gttgagtttc
     3601 tccagaagcg ttaatgtctg gcttctgata aagcgggcca tgttaagggc ggttttttcc
     3661 tgtttggtca ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt aatgataccg
     3721 atgaaacgag agaggatgct cacgatacgg gttactgatg atgaacatgc ccggttactg
     3781 gaacgttgtg agggtaaaca actggcggta tggatgcggc gggaccagag aaaaatcact
     3841 cagggtcaat gccagcgctt cgttaataca gatgtaggtg ttccacaggg tagccagcag
     3901 catcctgcga tgcagatccg gaacataatg gtgcagggcg ctgacttccg cgtttccaga
     3961 ctttacgaaa cacggaaacc gaagaccatt catgttgttg ctcaggtcgc agacgttttg
     4021 cagcagcagt cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta accagtaagg
     4081 caaccccgcc agcctagccg ggtcctcaac gacaggagca cgatcatgcg cacccgtggg
     4141 gccgccatgc cggcgataat ggcctgcttc tcgccgaaac gtttggtggc gggaccagtg
     4201 acgaaggctt gagcgagggc gtgcaagatt ccgaataccg caagcgacag gccgatcatc
     4261 gtcgcgctcc agcgaaagcg gtcctcgccg aaaatgaccc agagcgctgc cggcacctgt
     4321 cctacgagtt gcatgataaa gaagacagtc ataagtgcgg cgacgatagt catgccccgc
     4381 gcccaccgga aggagctgac tgggttgaag gctctcaagg gcatcggtcg agatcccggt
     4441 gcctaatgag tgagctaact tacattaatt gcgttgcgct cactgcccgc tttccagtcg
     4501 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
     4561 cgtattgggc gccagggtgg tttttctttt caccagtgag acgggcaaca gctgattgcc
     4621 cttcaccgcc tggccctgag agagttgcag caagcggtcc acgctggttt gccccagcag
     4681 gcgaaaatcc tgtttgatgg tggttaacgg cgggatataa catgagctgt cttcggtatc
     4741 gtcgtatccc actaccgaga tatccgcacc aacgcgcagc ccggactcgg taatggcgcg
     4801 cattgcgccc agcgccatct gatcgttggc aaccagcatc gcagtgggaa cgatgccctc
     4861 attcagcatt tgcatggttt gttgaaaacc ggacatggca ctccagtcgc cttcccgttc
     4921 cgctatcggc tgaatttgat tgcgagtgag atatttatgc cagccagcca gacgcagacg
     4981 cgccgagaca gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac ccaatgcgac
     5041 cagatgctcc acgcccagtc gcgtaccgtc ttcatgggag aaaataatac tgttgatggg
     5101 tgtctggtca gagacatcaa gaaataacgc cggaacatta gtgcaggcag cttccacagc
     5161 aatggcatcc tggtcatcca gcggatagtt aatgatcagc ccactgacgc gttgcgcgag
     5221 aagattgtgc accgccgctt tacaggcttc gacgccgctt cgttctacca tcgacaccac
     5281 cacgctggca cccagttgat cggcgcgaga tttaatcgcc gcgacaattt gcgacggcgc
     5341 gtgcagggcc agactggagg tggcaacgcc aatcagcaac gactgtttgc ccgccagttg
     5401 ttgtgccacg cggttgggaa tgtaattcag ctccgccatc gccgcttcca ctttttcccg
     5461 cgttttcgca gaaacgtggc tggcctggtt caccacgcgg gaaacggtct gataagagac
     5521 accggcatac tctgcgacat cgtataacgt tactggtttc acattcacca ccctgaattg
     5581 actctcttcc gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt cgatggtgtc
     5641 cgggatctcg acgctctccc ttatgcgact cctgcattag gaagcagccc agtagtaggt
     5701 tgaggccgtt gagcaccgcc gccgcaagga atggtgcatg caaggagatg gcgcccaaca
     5761 gtcccccggc cacggggcct gccaccatac ccacgccgaa acaagcgctc atgagcccga
     5821 agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca gcaaccgcac
     5881 ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta gaggatcgag atctcgatcc
     5941 cgcgaaatta atacgactca ctatagggga attgtgagcg gataacaatt cc
//
