LOCUS       sequence-364479-        5918 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for inducible E coli expression of the AP10 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_AP10-ehaA_LacI
FEATURES             Location/Qualifiers
     source          1..5918
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(6..20,21..26,27..43,44..50)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    26..42
                     /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    58..74
                     /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             93..104
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     sig_peptide     111..176
                     /codon_start=1
                     /gene="pelB (fragment)"
                     /product="leader peptide for secretion"
                     /label=pelB signal sequence
                     /translation="MKYLLPTAAAGLLLLAAQPAMA"
     terminator      1371..1465
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             1509..2168
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1575..1594)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      2233..2319
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(2412..3494)
                     /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     3456..3475
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3495..3572)
                     /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(3656..3678)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3764..3904
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3819..3838
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3919..3936)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4090..4678)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4170..4189)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4849..5640,5641..5709))
                     /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     5472..5491
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5710..5814)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5882..5900
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 agaaatcata aaaaatttat ttgctttgtg agcggataac aattataata gattcaattg
       61 tgagcggata acaatttcac acagaattca ttaaagagga gaaattaact atgaaatacc
      121 tgctgccgac cgctgctgct ggtctgctgc tcctcgctgc ccagccggcg atggccatga
      181 gaggatcgtc tccggaagac aagctggctc agattaagga aaagttgcaa caaattaaag
      241 aggaactggc cgcgaacgag gagaaattgc aggctaacaa atatggaggt agcggtagcg
      301 gctctggatc aggcagcgga tcactcgaga cacctacgcc gggtccggat ctgaatgtgg
      361 ataatgacct gcgaccggag gcgggtagct acattgcgaa ccttgcagca gcgaatacca
      421 tgttcaccac gcgtctgcat gagcgtctgg gtaatacgta ctataccgac atggtgacgg
      481 gtgagcagaa acaaaccact atgtggatgc gccatgaagg tggtcataat aaatggcgtg
      541 atggcagcgg ccagctgaaa acccaaagca atcgctatgt tctgcaactg ggaggcgatg
      601 tcgcgcagtg gagccaaaac ggcagcgacc gctggcatgt tggggtcatg gcgggatatg
      661 gcaacagcga cagcaaaacc atttcctcgc gaaccggtta tcgtgcaaaa gcgagtgtga
      721 acggatatag cacaggcctc tatgccacct ggtatgccga tgacgagtcg cgtaatggcg
      781 cgtatctcga cagttgggcg cagtacagct ggtttgataa cacagtgaaa ggggatgact
      841 tacaaagtga atcctataaa tcaaaaggat ttaccgcttc actggaagct ggatacaaac
      901 acaaattagc tgaatttaat ggcagccagg gaacgcgtaa tgaatggtat gttcagccgc
      961 aagcacaggt tacctggatg ggagtcaaag ccgataagca ccgcgaaagc aacggaaccc
     1021 tcgttcatag caacggtgat ggcaatgttc aaacccgact tggcgtaaaa acctggctga
     1081 agagccacca taaaatggat gacggtaaat cccgcgagtt ccagccgttt gtagaagtga
     1141 actggctaca taacagtaag gatttcagca ccagtatgga tggcgtgtct gtcactcagg
     1201 atggagcccg aaatattgct gagataaaaa ccggggtgga aggacagcta aatgccaacc
     1261 tgaatgtctg ggggaatgtg ggcgttcagg ttgccgatag gggatataat gacacctctg
     1321 caatggttgg cattaagtgg caattctgaa agcttaatta gctgagcttg gactcctgtt
     1381 gatagatcca gtaatgacct cagaactcca tctggatttg ttcagaacgc tcggttgccg
     1441 ccgggcgttt tttattggtg agaatccaag ctagcttggc gagattttca ggagctaagg
     1501 aagctaaaat ggagaaaaaa atcactggat ataccaccgt tgatatatcc caatggcatc
     1561 gtaaagaaca ttttgaggca tttcagtcag ttgctcaatg tacctataac cagaccgttc
     1621 agctggatat tacggccttt ttaaagaccg taaagaaaaa taagcacaag ttttatccgg
     1681 cctttattca cattcttgcc cgcctgatga atgctcatcc ggaatttcgt atggcaatga
     1741 aagacggtga gctggtgata tgggatagtg ttcacccttg ttacaccgtt ttccatgagc
     1801 aaactgaaac gttttcatcg ctctggagtg aataccacga cgatttccgg cagtttctac
     1861 acatatattc gcaagatgtg gcgtgttacg gtgaaaacct ggcctatttc cctaaagggt
     1921 ttattgagaa tatgtttttc gtctcagcca atccctgggt gagtttcacc agttttgatt
     1981 taaacgtggc caatatggac aacttcttcg cccccgtttt caccatgggc aaatattata
     2041 cgcaaggcga caaggtgctg atgccgctgg cgattcaggt tcatcatgcc gtttgtgatg
     2101 gcttccatgt cggcagaatg cttaatgaat tacaacagta ctgcgatgag tggcagggcg
     2161 gggcgtaatt tttttaaggc agttattggt gcccttaaac gcctggggta atgactctct
     2221 agcttgaggc atcaaataaa acgaaaggct cagtcgaaag actgggcctt tcgttttatc
     2281 tgttgtttgt cggtgaacgc tctcctgagt aggacaaatc cgccctctag attacgtgca
     2341 gtcgatgata agctgtcaaa catgagaatt gtgcctaatg agtgagctaa cttacattaa
     2401 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     2461 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgccagggt ggtttttctt
     2521 ttcaccagtg agacgggcaa cagctgattg cccttcaccg cctggccctg agagagttgc
     2581 agcaagcggt ccacgctggt ttgccccagc aggcgaaaat cctgtttgat ggtggttaac
     2641 ggcgggatat aacatgagct gtcttcggta tcgtcgtatc ccactaccga gatatccgca
     2701 ccaacgcgca gcccggactc ggtaatggcg cgcattgcgc ccagcgccat ctgatcgttg
     2761 gcaaccagca tcgcagtggg aacgatgccc tcattcagca tttgcatggt ttgttgaaaa
     2821 ccggacatgg cactccagtc gccttcccgt tccgctatcg gctgaatttg attgcgagtg
     2881 agatatttat gccagccagc cagacgcaga cgcgccgaga cagaacttaa tgggcccgct
     2941 aacagcgcga tttgctggtg acccaatgcg accagatgct ccacgcccag tcgcgtaccg
     3001 tcttcatggg agaaaataat actgttgatg ggtgtctggt cagagacatc aagaaataac
     3061 gccggaacat tagtgcaggc agcttccaca gcaatggcat cctggtcatc cagcggatag
     3121 ttaatgatca gcccactgac gcgttgcgcg agaagattgt gcaccgccgc tttacaggct
     3181 tcgacgccgc ttcgttctac catcgacacc accacgctgg cacccagttg atcggcgcga
     3241 gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga ggtggcaacg
     3301 ccaatcagca acgactgttt gcccgccagt tgttgtgcca cgcggttggg aatgtaattc
     3361 agctccgcca tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg gctggcctgg
     3421 ttcaccacgc gggaaacggt ctgataagag acaccggcat actctgcgac atcgtataac
     3481 gttactggtt tcacattcac caccctgaat tgactctctt ccgggcgcta tcatgccata
     3541 ccgcgaaagg ttttgcacca ttcgatggtg tcggaatttc gggcagcgtt gggtcctggc
     3601 cacgggtgcg catgatctag agctgcctcg cgcgtttcgg tgatgacggt gaaaacctct
     3661 gacacatgca gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac
     3721 aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg gggcgcagcc atgacccagt
     3781 cacgtagcga tagcggagtg tatactggct taactatgcg gcatcagagc agattgtact
     3841 gagagtgcac catatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat
     3901 caggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     3961 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     4021 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     4081 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     4141 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     4201 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     4261 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     4321 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     4381 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     4441 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     4501 gtggtggcct aactacggct acactagaag gacagtattt ggtatctgcg ctctgctgaa
     4561 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     4621 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     4681 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     4741 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     4801 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     4861 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     4921 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     4981 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     5041 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     5101 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     5161 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     5221 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     5281 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     5341 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     5401 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     5461 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     5521 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     5581 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     5641 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     5701 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     5761 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     5821 tccccgaaaa gtgccacctg acgtctaaga aaccattatt atcatgacat taacctataa
     5881 aaataggcgt atcacgaggc cctttcgtct tcacgtcg
//
