LOCUS       sequence-345149-        5884 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  gfcB protein expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5884)
  AUTHORS   Larson MR, Biddle K, Gorman A, Boutom S, Rosenshine I, Saper MA
  TITLE     Escherichia coli O127 group 4 capsule proteins assemble at the outer
            membrane.
  JOURNAL   PLoS One. 2021 Nov 15;16(11):e0259900. doi: 
            10.1371/journal.pone.0259900. eCollection 2021.
  PUBMED    34780538
REFERENCE   2  (bases 1 to 5884)
  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: "PLoS One.";
            date: "2021-11-15"; volume: "16(11):e0259900. doi"; pages: " 
            10.1371/journal.pone.0259900. eCollection 2021"
COMMENT     Sequence Label: pMCSG7(gfcB)
FEATURES             Location/Qualifiers
     source          1..5884
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1299..1490
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(1484..1506)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    1592..1731
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     1647..1666
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(1746..1763)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1917..2505)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1997..2016)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2676..3467,3468..3536))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3299..3318
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3537..3640)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3807..3826
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(3950..3997)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     3993..4011
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(4064..4081)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(4748..4768)
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             complement(4793..4810)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             4821..4843
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    4858..4882
                     /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)."
     primer_bind     complement(4882..4901)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4883..4901)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4989..5008)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        5214..5291
                     /gene="lacI"
                     /label=lacI promoter
     CDS             join(5292..5884,1..490)
                     /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     complement(5311..5330)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 ggctggctgg cataaatatc tcactcgcaa tcaaattcag ccgatagcgg aacgggaagg
       61 cgactggagt gccatgtccg gttttcaaca aaccatgcaa atgctgaatg agggcatcgt
      121 tcccactgcg atgctggttg ccaacgatca gatggcgctg ggcgcaatgc gcgccattac
      181 cgagtccggg ctgcgcgttg gtgcggatat ctcggtagtg ggatacgacg ataccgaaga
      241 cagctcatgt tatatcccgc cgttaaccac catcaaacag gattttcgcc tgctggggca
      301 aaccagcgtg gaccgcttgc tgcaactctc tcagggccag gcggtgaagg gcaatcagct
      361 gttgcccgtc tcactggtga aaagaaaaac caccctggcg cccaatacgc aaaccgcctc
      421 tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag
      481 cgggcagtga gcgcaacgca attaatgtaa gttagctcac tcattaggca ccgggatctc
      541 gaccgatgcc cttgagagcc ttcaacccag tcagctcctt ccggtgggcg cggggcatga
      601 ctatcgtcgc cgcacttatg actgtcttct ttatcatgca actcgtagga caggtgccgg
      661 cagcgctctg ggtcattttc ggcgaggacc gctttcgctg gagcgcgacg atgatcggcc
      721 tgtcgcttgc ggtattcgga atcttgcacg ccctcgctca agccttcgtc actggtcccg
      781 ccaccaaacg tttcggcgag aagcaggcca ttatcgccgg catggcggcc ccacgggtgc
      841 gcatgatcgt gctcctgtcg ttgaggaccc ggctaggctg gcggggttgc cttactggtt
      901 agcagaatga atcaccgata cgcgagcgaa cgtgaagcga ctgctgctgc aaaacgtctg
      961 cgacctgagc aacaacatga atggtcttcg gtttccgtgt ttcgtaaagt ctggaaacgc
     1021 ggaagtcagc gccctgcacc attatgttcc ggatctgcat cgcaggatgc tgctggctac
     1081 cctgtggaac acctacatct gtattaacga agcgctggca ttgaccctga gtgatttttc
     1141 tctggtcccg ccgcatccat accgccagtt gtttaccctc acaacgttcc agtaaccggg
     1201 catgttcatc atcagtaacc cgtatcgtga gcatcctctc tcgtttcatc ggtatcatta
     1261 cccccatgaa cagaaatccc ccttacacgg aggcatcagt gaccaaacag gaaaaaaccg
     1321 cccttaacat ggcccgcttt atcagaagcc agacattaac gcttctggag aaactcaacg
     1381 agctggacgc ggatgaacag gcagacatct gtgaatcgct tcacgaccac gctgatgagc
     1441 tttaccgcag ctgcctcgcg cgtttcggtg atgacggtga aaacctctga cacatgcagc
     1501 tcccggagac ggtcacagct tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg
     1561 gcgcgtcagc gggtgttggc gggtgtcggg gcgcagccat gacccagtca cgtagcgata
     1621 gcggagtgta tactggctta actatgcggc atcagagcag attgtactga gagtgcacca
     1681 ttgcggtgtg aaataccgca cagatgcgta aggagaaaat accgcatcag gcgctcttcc
     1741 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
     1801 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     1861 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     1921 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     1981 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     2041 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     2101 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     2161 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     2221 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     2281 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
     2341 tacggctaca ctagaaggac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     2401 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     2461 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     2521 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
     2581 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
     2641 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
     2701 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
     2761 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
     2821 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
     2881 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
     2941 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tgcaggcatc
     3001 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
     3061 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
     3121 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
     3181 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
     3241 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
     3301 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     3361 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     3421 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     3481 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     3541 ttcctttttc aaattattga agcatttatc agggttattg tctcatgagc ggatacatat
     3601 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     3661 cacctaaatt gtaagcgtta atgtgaacca tcaccctaat caagtttttt ggggtcgagg
     3721 tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga
     3781 aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg
     3841 ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg
     3901 ctacagggcg cgtcccattc gccaatccgg atatagttcc tcctttcagc aaaaaacccc
     3961 tcaagacccg tttagaggcc ccaaggggtt atgctagtta ttgctcagcg gtggcagcag
     4021 ccaactcagc ttcctttcgg gctttgttag cagccggatc tcagtggtgg tggtggtggt
     4081 gctcgagtgc ggccgcaagc ttgtcgacgg agctcgaatt cggatccgtt atccacttcc
     4141 aatgtcatgg tgcgggttta agaaacgtca tttctacggg aataacccct gcgcctaaca
     4201 tttgccgact ttgacgaacc tggccagtag ccgaatcgac ccagaatgtg ttttgccact
     4261 gtttttctgg cagcgtggaa gtcacctctt cctgccagac attacacagc acatcgcgac
     4321 ccgcaatatt cagcgtctct aatccagcaa aggaaaaagt ggaactcagg gtcgcagagc
     4381 gaaaatggtt gtcttcgctc cagagaatat cgcgcgtcca gcggctgccg tcttttattg
     4441 ccagcgcgtt acgcagaggg tcctgcccgg aatttgtcac ttccagcaga ttattattaa
     4501 gtttgacggt ttttaaaaga cgtccattgt gggtaaccag catggcgtta tcctgggata
     4561 accatttgct ttgttcttgt tctatatatc ccagtaccac aaagattcgt tgcccttcat
     4621 taaggcgtaa atacatcgtg gaataaggca acgcctggat ctgctgatcc gctacggtga
     4681 tatcctgatt atcgaatagg ctggcgcgaa atgtatcgac catactttgc tggctgtgcg
     4741 tagcattgga ttggaagtac aggttctcgg tacccagatc tacaccagaa gaatgatgat
     4801 gatgatggtg catatgtata tctccttctt aaagttaaac aaaattattt ctagagggga
     4861 attgttatcc gctcacaatt cccctatagt gagtcgtatt aatttcgcgg gatcgagatc
     4921 gatctcgatc ctctacgccg gacgcatcgt ggccggcatc accggcgcca caggtgcggt
     4981 tgctggcgcc tatatcgccg acatcaccga tggggaagat cgggctcgcc acttcgggct
     5041 catgagcgct tgtttcggcg tgggtatggt ggcaggcccc gtggccgggg gactgttggg
     5101 cgccatctcc ttgcatgcac cattccttgc ggcggcggtg ctcaacggcc tcaacctact
     5161 actgggctgc ttcctaatgc aggagtcgca taagggagag cgtcgagatc ccggacacca
     5221 tcgaatggcg caaaaccttt cgcggtatgg catgatagcg cccggaagag agtcaattca
     5281 gggtggtgaa tgtgaaacca gtaacgttat acgatgtcgc agagtatgcc ggtgtctctt
     5341 atcagaccgt ttcccgcgtg gtgaaccagg ccagccacgt ttctgcgaaa acgcgggaaa
     5401 aagtggaagc ggcgatggcg gagctgaatt acattcccaa ccgcgtggca caacaactgg
     5461 cgggcaaaca gtcgttgctg attggcgttg ccacctccag tctggccctg cacgcgccgt
     5521 cgcaaattgt cgcggcgatt aaatctcgcg ccgatcaact gggtgccagc gtggtggtgt
     5581 cgatggtaga acgaagcggc gtcgaagcct gtaaagcggc ggtgcacaat cttctcgcgc
     5641 aacgcgtcag tgggctgatc attaactatc cgctggatga ccaggatgcc attgctgtgg
     5701 aagctgcctg cactaatgtt ccggcgttat ttcttgatgt ctctgaccag acacccatca
     5761 acagtattat tttctcccat gaagacggta cgcgactggg cgtggagcat ctggtcgcat
     5821 tgggtcacca gcaaatcgcg ctgttagcgg gcccattaag ttctgtctcg gcgcgtctgc
     5881 gtct
//
