LOCUS       sequence-366946-        5918 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for inducible E coli expression of the P9 helixCAM..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5918)
  AUTHORS   Chao G, Wannier TM, Gutierrez C, Borders NC, Appleton E, Chadha A, 
            Lebar T, Church GM
  TITLE     helixCAM: A platform for programmable cellular assembly in bacteria 
            and human cells.
  JOURNAL   Cell. 2022 Aug 23. pii: S0092-8674(22)01061-3. doi: 
            10.1016/j.cell.2022.08.012.
  PUBMED    36055250
REFERENCE   2  (bases 1 to 5918)
  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: "Cell. 2022 
            Aug 23. pii: S0092-8674(22)01061-3. doi: 
            10.1016/j.cell.2022.08.012."
COMMENT     Sequence Label: pQE80_T5_P9-ehaA_LacI
FEATURES             Location/Qualifiers
     source          1..5918
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(10..24,25..30,31..47,48..54)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    30..46
                     /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)."
     protein_bind    62..78
                     /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             97..108
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     sig_peptide     115..180
                     /codon_start=1
                     /gene="pelB (fragment)"
                     /product="leader peptide for secretion"
                     /label=pelB signal sequence
                     /translation="MKYLLPTAAAGLLLLAAQPAMA"
     terminator      1375..1469
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             1513..2172
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1579..1598)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      2237..2323
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(2416..3498)
                     /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     3460..3479
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3499..3576)
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     primer_bind     complement(3660..3682)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3768..3908
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3823..3842
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3923..3940)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4094..4682)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4174..4193)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4853..5644,5645..5713))
                     /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     5476..5495
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5714..5818)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5886..5904
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 gtcgagaaat cataaaaaat ttatttgctt tgtgagcgga taacaattat aatagattca
       61 attgtgagcg gataacaatt tcacacagaa ttcattaaag aggagaaatt aactatgaaa
      121 tacctgctgc cgaccgctgc tgctggtctg ctgctcctcg ctgcccagcc ggcgatggcc
      181 atgagaggat cgtctcccga agatgagaat caggcgcttg aacagaagaa tgcccaactg
      241 aagcaggaaa ttgccgccct ggaacaggag attgcacaat tggagtacgg aggtagcggt
      301 agcggctctg gatcaggcag cggatcactc gagacaccta cgccgggtcc ggatctgaat
      361 gtggataatg acctgcgacc ggaggcgggt agctacattg cgaaccttgc agcagcgaat
      421 accatgttca ccacgcgtct gcatgagcgt ctgggtaata cgtactatac cgacatggtg
      481 acgggtgagc agaaacaaac cactatgtgg atgcgccatg aaggtggtca taataaatgg
      541 cgtgatggca gcggccagct gaaaacccaa agcaatcgct atgttctgca actgggaggc
      601 gatgtcgcgc agtggagcca aaacggcagc gaccgctggc atgttggggt catggcggga
      661 tatggcaaca gcgacagcaa aaccatttcc tcgcgaaccg gttatcgtgc aaaagcgagt
      721 gtgaacggat atagcacagg cctctatgcc acctggtatg ccgatgacga gtcgcgtaat
      781 ggcgcgtatc tcgacagttg ggcgcagtac agctggtttg ataacacagt gaaaggggat
      841 gacttacaaa gtgaatccta taaatcaaaa ggatttaccg cttcactgga agctggatac
      901 aaacacaaat tagctgaatt taatggcagc cagggaacgc gtaatgaatg gtatgttcag
      961 ccgcaagcac aggttacctg gatgggagtc aaagccgata agcaccgcga aagcaacgga
     1021 accctcgttc atagcaacgg tgatggcaat gttcaaaccc gacttggcgt aaaaacctgg
     1081 ctgaagagcc accataaaat ggatgacggt aaatcccgcg agttccagcc gtttgtagaa
     1141 gtgaactggc tacataacag taaggatttc agcaccagta tggatggcgt gtctgtcact
     1201 caggatggag cccgaaatat tgctgagata aaaaccgggg tggaaggaca gctaaatgcc
     1261 aacctgaatg tctgggggaa tgtgggcgtt caggttgccg ataggggata taatgacacc
     1321 tctgcaatgg ttggcattaa gtggcaattc tgaaagctta attagctgag cttggactcc
     1381 tgttgataga tccagtaatg acctcagaac tccatctgga tttgttcaga acgctcggtt
     1441 gccgccgggc gttttttatt ggtgagaatc caagctagct tggcgagatt ttcaggagct
     1501 aaggaagcta aaatggagaa aaaaatcact ggatatacca ccgttgatat atcccaatgg
     1561 catcgtaaag aacattttga ggcatttcag tcagttgctc aatgtaccta taaccagacc
     1621 gttcagctgg atattacggc ctttttaaag accgtaaaga aaaataagca caagttttat
     1681 ccggccttta ttcacattct tgcccgcctg atgaatgctc atccggaatt tcgtatggca
     1741 atgaaagacg gtgagctggt gatatgggat agtgttcacc cttgttacac cgttttccat
     1801 gagcaaactg aaacgttttc atcgctctgg agtgaatacc acgacgattt ccggcagttt
     1861 ctacacatat attcgcaaga tgtggcgtgt tacggtgaaa acctggccta tttccctaaa
     1921 gggtttattg agaatatgtt tttcgtctca gccaatccct gggtgagttt caccagtttt
     1981 gatttaaacg tggccaatat ggacaacttc ttcgcccccg ttttcaccat gggcaaatat
     2041 tatacgcaag gcgacaaggt gctgatgccg ctggcgattc aggttcatca tgccgtttgt
     2101 gatggcttcc atgtcggcag aatgcttaat gaattacaac agtactgcga tgagtggcag
     2161 ggcggggcgt aattttttta aggcagttat tggtgccctt aaacgcctgg ggtaatgact
     2221 ctctagcttg aggcatcaaa taaaacgaaa ggctcagtcg aaagactggg cctttcgttt
     2281 tatctgttgt ttgtcggtga acgctctcct gagtaggaca aatccgccct ctagattacg
     2341 tgcagtcgat gataagctgt caaacatgag aattgtgcct aatgagtgag ctaacttaca
     2401 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     2461 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgcca gggtggtttt
     2521 tcttttcacc agtgagacgg gcaacagctg attgcccttc accgcctggc cctgagagag
     2581 ttgcagcaag cggtccacgc tggtttgccc cagcaggcga aaatcctgtt tgatggtggt
     2641 taacggcggg atataacatg agctgtcttc ggtatcgtcg tatcccacta ccgagatatc
     2701 cgcaccaacg cgcagcccgg actcggtaat ggcgcgcatt gcgcccagcg ccatctgatc
     2761 gttggcaacc agcatcgcag tgggaacgat gccctcattc agcatttgca tggtttgttg
     2821 aaaaccggac atggcactcc agtcgccttc ccgttccgct atcggctgaa tttgattgcg
     2881 agtgagatat ttatgccagc cagccagacg cagacgcgcc gagacagaac ttaatgggcc
     2941 cgctaacagc gcgatttgct ggtgacccaa tgcgaccaga tgctccacgc ccagtcgcgt
     3001 accgtcttca tgggagaaaa taatactgtt gatgggtgtc tggtcagaga catcaagaaa
     3061 taacgccgga acattagtgc aggcagcttc cacagcaatg gcatcctggt catccagcgg
     3121 atagttaatg atcagcccac tgacgcgttg cgcgagaaga ttgtgcaccg ccgctttaca
     3181 ggcttcgacg ccgcttcgtt ctaccatcga caccaccacg ctggcaccca gttgatcggc
     3241 gcgagattta atcgccgcga caatttgcga cggcgcgtgc agggccagac tggaggtggc
     3301 aacgccaatc agcaacgact gtttgcccgc cagttgttgt gccacgcggt tgggaatgta
     3361 attcagctcc gccatcgccg cttccacttt ttcccgcgtt ttcgcagaaa cgtggctggc
     3421 ctggttcacc acgcgggaaa cggtctgata agagacaccg gcatactctg cgacatcgta
     3481 taacgttact ggtttcacat tcaccaccct gaattgactc tcttccgggc gctatcatgc
     3541 cataccgcga aaggttttgc accattcgat ggtgtcggaa tttcgggcag cgttgggtcc
     3601 tggccacggg tgcgcatgat ctagagctgc ctcgcgcgtt tcggtgatga cggtgaaaac
     3661 ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc
     3721 agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggcgc agccatgacc
     3781 cagtcacgta gcgatagcgg agtgtatact ggcttaacta tgcggcatca gagcagattg
     3841 tactgagagt gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc
     3901 gcatcaggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     3961 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     4021 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     4081 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     4141 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     4201 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     4261 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     4321 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     4381 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     4441 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     4501 tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc
     4561 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     4621 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     4681 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     4741 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     4801 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     4861 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     4921 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     4981 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     5041 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     5101 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     5161 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     5221 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     5281 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     5341 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     5401 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     5461 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     5521 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     5581 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     5641 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     5701 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     5761 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     5821 catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct
     5881 ataaaaatag gcgtatcacg aggccctttc gtcttcac
//
