LOCUS       sequence-366933-        5918 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for inducible E coli expression of the AP4 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_AP4-ehaA_LacI
FEATURES             Location/Qualifiers
     source          1..5918
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     43..61
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(85..99,100..105,106..122,123..129)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    105..121
                     /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    137..153
                     /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             172..183
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     sig_peptide     190..255
                     /codon_start=1
                     /gene="pelB (fragment)"
                     /product="leader peptide for secretion"
                     /label=pelB signal sequence
                     /translation="MKYLLPTAAAGLLLLAAQPAMA"
     terminator      1450..1544
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             1588..2247
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1654..1673)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      2312..2398
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(2491..3573)
                     /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     3535..3554
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3574..3651)
                     /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(3735..3757)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3843..3983
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3898..3917
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3998..4015)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4169..4757)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4249..4268)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4928..5719,5720..5788))
                     /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     5551..5570
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5789..5893)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg tatcacgagg
       61 ccctttcgtc ttcacgtcga gaaatcataa aaaatttatt tgctttgtga gcggataaca
      121 attataatag attcaattgt gagcggataa caatttcaca cagaattcat taaagaggag
      181 aaattaacta tgaaatacct gctgccgacc gctgctgctg gtctgctgct cctcgctgcc
      241 cagccggcga tggccatgag aggatcgtct cccgaggatg aattagcagc taatgaagag
      301 gaactgcaac agaacgaaca aaagctggct caaattaaac agaagttgca ggctatcaag
      361 tacggaggta gcggtagcgg ctctggatca ggcagcggat cactcgagac acctacgccg
      421 ggtccggatc tgaatgtgga taatgacctg cgaccggagg cgggtagcta cattgcgaac
      481 cttgcagcag cgaataccat gttcaccacg cgtctgcatg agcgtctggg taatacgtac
      541 tataccgaca tggtgacggg tgagcagaaa caaaccacta tgtggatgcg ccatgaaggt
      601 ggtcataata aatggcgtga tggcagcggc cagctgaaaa cccaaagcaa tcgctatgtt
      661 ctgcaactgg gaggcgatgt cgcgcagtgg agccaaaacg gcagcgaccg ctggcatgtt
      721 ggggtcatgg cgggatatgg caacagcgac agcaaaacca tttcctcgcg aaccggttat
      781 cgtgcaaaag cgagtgtgaa cggatatagc acaggcctct atgccacctg gtatgccgat
      841 gacgagtcgc gtaatggcgc gtatctcgac agttgggcgc agtacagctg gtttgataac
      901 acagtgaaag gggatgactt acaaagtgaa tcctataaat caaaaggatt taccgcttca
      961 ctggaagctg gatacaaaca caaattagct gaatttaatg gcagccaggg aacgcgtaat
     1021 gaatggtatg ttcagccgca agcacaggtt acctggatgg gagtcaaagc cgataagcac
     1081 cgcgaaagca acggaaccct cgttcatagc aacggtgatg gcaatgttca aacccgactt
     1141 ggcgtaaaaa cctggctgaa gagccaccat aaaatggatg acggtaaatc ccgcgagttc
     1201 cagccgtttg tagaagtgaa ctggctacat aacagtaagg atttcagcac cagtatggat
     1261 ggcgtgtctg tcactcagga tggagcccga aatattgctg agataaaaac cggggtggaa
     1321 ggacagctaa atgccaacct gaatgtctgg gggaatgtgg gcgttcaggt tgccgatagg
     1381 ggatataatg acacctctgc aatggttggc attaagtggc aattctgaaa gcttaattag
     1441 ctgagcttgg actcctgttg atagatccag taatgacctc agaactccat ctggatttgt
     1501 tcagaacgct cggttgccgc cgggcgtttt ttattggtga gaatccaagc tagcttggcg
     1561 agattttcag gagctaagga agctaaaatg gagaaaaaaa tcactggata taccaccgtt
     1621 gatatatccc aatggcatcg taaagaacat tttgaggcat ttcagtcagt tgctcaatgt
     1681 acctataacc agaccgttca gctggatatt acggcctttt taaagaccgt aaagaaaaat
     1741 aagcacaagt tttatccggc ctttattcac attcttgccc gcctgatgaa tgctcatccg
     1801 gaatttcgta tggcaatgaa agacggtgag ctggtgatat gggatagtgt tcacccttgt
     1861 tacaccgttt tccatgagca aactgaaacg ttttcatcgc tctggagtga ataccacgac
     1921 gatttccggc agtttctaca catatattcg caagatgtgg cgtgttacgg tgaaaacctg
     1981 gcctatttcc ctaaagggtt tattgagaat atgtttttcg tctcagccaa tccctgggtg
     2041 agtttcacca gttttgattt aaacgtggcc aatatggaca acttcttcgc ccccgttttc
     2101 accatgggca aatattatac gcaaggcgac aaggtgctga tgccgctggc gattcaggtt
     2161 catcatgccg tttgtgatgg cttccatgtc ggcagaatgc ttaatgaatt acaacagtac
     2221 tgcgatgagt ggcagggcgg ggcgtaattt ttttaaggca gttattggtg cccttaaacg
     2281 cctggggtaa tgactctcta gcttgaggca tcaaataaaa cgaaaggctc agtcgaaaga
     2341 ctgggccttt cgttttatct gttgtttgtc ggtgaacgct ctcctgagta ggacaaatcc
     2401 gccctctaga ttacgtgcag tcgatgataa gctgtcaaac atgagaattg tgcctaatga
     2461 gtgagctaac ttacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     2521 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     2581 cgccagggtg gtttttcttt tcaccagtga gacgggcaac agctgattgc ccttcaccgc
     2641 ctggccctga gagagttgca gcaagcggtc cacgctggtt tgccccagca ggcgaaaatc
     2701 ctgtttgatg gtggttaacg gcgggatata acatgagctg tcttcggtat cgtcgtatcc
     2761 cactaccgag atatccgcac caacgcgcag cccggactcg gtaatggcgc gcattgcgcc
     2821 cagcgccatc tgatcgttgg caaccagcat cgcagtggga acgatgccct cattcagcat
     2881 ttgcatggtt tgttgaaaac cggacatggc actccagtcg ccttcccgtt ccgctatcgg
     2941 ctgaatttga ttgcgagtga gatatttatg ccagccagcc agacgcagac gcgccgagac
     3001 agaacttaat gggcccgcta acagcgcgat ttgctggtga cccaatgcga ccagatgctc
     3061 cacgcccagt cgcgtaccgt cttcatggga gaaaataata ctgttgatgg gtgtctggtc
     3121 agagacatca agaaataacg ccggaacatt agtgcaggca gcttccacag caatggcatc
     3181 ctggtcatcc agcggatagt taatgatcag cccactgacg cgttgcgcga gaagattgtg
     3241 caccgccgct ttacaggctt cgacgccgct tcgttctacc atcgacacca ccacgctggc
     3301 acccagttga tcggcgcgag atttaatcgc cgcgacaatt tgcgacggcg cgtgcagggc
     3361 cagactggag gtggcaacgc caatcagcaa cgactgtttg cccgccagtt gttgtgccac
     3421 gcggttggga atgtaattca gctccgccat cgccgcttcc actttttccc gcgttttcgc
     3481 agaaacgtgg ctggcctggt tcaccacgcg ggaaacggtc tgataagaga caccggcata
     3541 ctctgcgaca tcgtataacg ttactggttt cacattcacc accctgaatt gactctcttc
     3601 cgggcgctat catgccatac cgcgaaaggt tttgcaccat tcgatggtgt cggaatttcg
     3661 ggcagcgttg ggtcctggcc acgggtgcgc atgatctaga gctgcctcgc gcgtttcggt
     3721 gatgacggtg aaaacctctg acacatgcag ctcccggaga cggtcacagc ttgtctgtaa
     3781 gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg
     3841 ggcgcagcca tgacccagtc acgtagcgat agcggagtgt atactggctt aactatgcgg
     3901 catcagagca gattgtactg agagtgcacc atatgcggtg tgaaataccg cacagatgcg
     3961 taaggagaaa ataccgcatc aggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     4021 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     4081 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     4141 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     4201 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     4261 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     4321 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     4381 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     4441 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     4501 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     4561 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg
     4621 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     4681 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     4741 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     4801 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     4861 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     4921 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     4981 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     5041 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     5101 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     5161 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     5221 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     5281 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     5341 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     5401 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     5461 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     5521 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     5581 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     5641 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     5701 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     5761 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     5821 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     5881 taggggttcc gcgcacattt ccccgaaaag tgccacct
//
