LOCUS       sequence-365756-        5918 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for inducible E coli expression of the P3 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_P3-ehaA_LacI
FEATURES             Location/Qualifiers
     source          1..5918
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     124..142
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(166..180,181..186,187..203,204..210)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    186..202
                     /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    218..234
                     /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             253..264
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     sig_peptide     271..336
                     /codon_start=1
                     /gene="pelB (fragment)"
                     /product="leader peptide for secretion"
                     /label=pelB signal sequence
                     /translation="MKYLLPTAAAGLLLLAAQPAMA"
     terminator      1531..1625
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             1669..2328
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1735..1754)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      2393..2479
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(2572..3654)
                     /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     3616..3635
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3655..3732)
                     /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(3816..3838)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3924..4064
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3979..3998
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4079..4096)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4250..4838)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4330..4349)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5009..5800,5801..5869))
                     /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     5632..5651
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(5870..5918,1..56))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
       61 tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac attaacctat
      121 aaaaataggc gtatcacgag gccctttcgt cttcacgtcg agaaatcata aaaaatttat
      181 ttgctttgtg agcggataac aattataata gattcaattg tgagcggata acaatttcac
      241 acagaattca ttaaagagga gaaattaact atgaaatacc tgctgccgac cgctgctgct
      301 ggtctgctgc tcctcgctgc ccagccggcg atggccatga gaggatcgtc gcctgaggac
      361 gaaatccaac aacttgagga ggagattgca caattggagc aaaaaaatgc tgcgctgaaa
      421 gagaaaaatc aggcgcttaa atacggcggt agcggtagcg gctctggatc aggcagcgga
      481 tcactcgaga cacctacgcc gggtccggat ctgaatgtgg ataatgacct gcgaccggag
      541 gcgggtagct acattgcgaa ccttgcagca gcgaatacca tgttcaccac gcgtctgcat
      601 gagcgtctgg gtaatacgta ctataccgac atggtgacgg gtgagcagaa acaaaccact
      661 atgtggatgc gccatgaagg tggtcataat aaatggcgtg atggcagcgg ccagctgaaa
      721 acccaaagca atcgctatgt tctgcaactg ggaggcgatg tcgcgcagtg gagccaaaac
      781 ggcagcgacc gctggcatgt tggggtcatg gcgggatatg gcaacagcga cagcaaaacc
      841 atttcctcgc gaaccggtta tcgtgcaaaa gcgagtgtga acggatatag cacaggcctc
      901 tatgccacct ggtatgccga tgacgagtcg cgtaatggcg cgtatctcga cagttgggcg
      961 cagtacagct ggtttgataa cacagtgaaa ggggatgact tacaaagtga atcctataaa
     1021 tcaaaaggat ttaccgcttc actggaagct ggatacaaac acaaattagc tgaatttaat
     1081 ggcagccagg gaacgcgtaa tgaatggtat gttcagccgc aagcacaggt tacctggatg
     1141 ggagtcaaag ccgataagca ccgcgaaagc aacggaaccc tcgttcatag caacggtgat
     1201 ggcaatgttc aaacccgact tggcgtaaaa acctggctga agagccacca taaaatggat
     1261 gacggtaaat cccgcgagtt ccagccgttt gtagaagtga actggctaca taacagtaag
     1321 gatttcagca ccagtatgga tggcgtgtct gtcactcagg atggagcccg aaatattgct
     1381 gagataaaaa ccggggtgga aggacagcta aatgccaacc tgaatgtctg ggggaatgtg
     1441 ggcgttcagg ttgccgatag gggatataat gacacctctg caatggttgg cattaagtgg
     1501 caattctgaa agcttaatta gctgagcttg gactcctgtt gatagatcca gtaatgacct
     1561 cagaactcca tctggatttg ttcagaacgc tcggttgccg ccgggcgttt tttattggtg
     1621 agaatccaag ctagcttggc gagattttca ggagctaagg aagctaaaat ggagaaaaaa
     1681 atcactggat ataccaccgt tgatatatcc caatggcatc gtaaagaaca ttttgaggca
     1741 tttcagtcag ttgctcaatg tacctataac cagaccgttc agctggatat tacggccttt
     1801 ttaaagaccg taaagaaaaa taagcacaag ttttatccgg cctttattca cattcttgcc
     1861 cgcctgatga atgctcatcc ggaatttcgt atggcaatga aagacggtga gctggtgata
     1921 tgggatagtg ttcacccttg ttacaccgtt ttccatgagc aaactgaaac gttttcatcg
     1981 ctctggagtg aataccacga cgatttccgg cagtttctac acatatattc gcaagatgtg
     2041 gcgtgttacg gtgaaaacct ggcctatttc cctaaagggt ttattgagaa tatgtttttc
     2101 gtctcagcca atccctgggt gagtttcacc agttttgatt taaacgtggc caatatggac
     2161 aacttcttcg cccccgtttt caccatgggc aaatattata cgcaaggcga caaggtgctg
     2221 atgccgctgg cgattcaggt tcatcatgcc gtttgtgatg gcttccatgt cggcagaatg
     2281 cttaatgaat tacaacagta ctgcgatgag tggcagggcg gggcgtaatt tttttaaggc
     2341 agttattggt gcccttaaac gcctggggta atgactctct agcttgaggc atcaaataaa
     2401 acgaaaggct cagtcgaaag actgggcctt tcgttttatc tgttgtttgt cggtgaacgc
     2461 tctcctgagt aggacaaatc cgccctctag attacgtgca gtcgatgata agctgtcaaa
     2521 catgagaatt gtgcctaatg agtgagctaa cttacattaa ttgcgttgcg ctcactgccc
     2581 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     2641 agaggcggtt tgcgtattgg gcgccagggt ggtttttctt ttcaccagtg agacgggcaa
     2701 cagctgattg cccttcaccg cctggccctg agagagttgc agcaagcggt ccacgctggt
     2761 ttgccccagc aggcgaaaat cctgtttgat ggtggttaac ggcgggatat aacatgagct
     2821 gtcttcggta tcgtcgtatc ccactaccga gatatccgca ccaacgcgca gcccggactc
     2881 ggtaatggcg cgcattgcgc ccagcgccat ctgatcgttg gcaaccagca tcgcagtggg
     2941 aacgatgccc tcattcagca tttgcatggt ttgttgaaaa ccggacatgg cactccagtc
     3001 gccttcccgt tccgctatcg gctgaatttg attgcgagtg agatatttat gccagccagc
     3061 cagacgcaga cgcgccgaga cagaacttaa tgggcccgct aacagcgcga tttgctggtg
     3121 acccaatgcg accagatgct ccacgcccag tcgcgtaccg tcttcatggg agaaaataat
     3181 actgttgatg ggtgtctggt cagagacatc aagaaataac gccggaacat tagtgcaggc
     3241 agcttccaca gcaatggcat cctggtcatc cagcggatag ttaatgatca gcccactgac
     3301 gcgttgcgcg agaagattgt gcaccgccgc tttacaggct tcgacgccgc ttcgttctac
     3361 catcgacacc accacgctgg cacccagttg atcggcgcga gatttaatcg ccgcgacaat
     3421 ttgcgacggc gcgtgcaggg ccagactgga ggtggcaacg ccaatcagca acgactgttt
     3481 gcccgccagt tgttgtgcca cgcggttggg aatgtaattc agctccgcca tcgccgcttc
     3541 cactttttcc cgcgttttcg cagaaacgtg gctggcctgg ttcaccacgc gggaaacggt
     3601 ctgataagag acaccggcat actctgcgac atcgtataac gttactggtt tcacattcac
     3661 caccctgaat tgactctctt ccgggcgcta tcatgccata ccgcgaaagg ttttgcacca
     3721 ttcgatggtg tcggaatttc gggcagcgtt gggtcctggc cacgggtgcg catgatctag
     3781 agctgcctcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca gctcccggag
     3841 acggtcacag cttgtctgta agcggatgcc gggagcagac aagcccgtca gggcgcgtca
     3901 gcgggtgttg gcgggtgtcg gggcgcagcc atgacccagt cacgtagcga tagcggagtg
     3961 tatactggct taactatgcg gcatcagagc agattgtact gagagtgcac catatgcggt
     4021 gtgaaatacc gcacagatgc gtaaggagaa aataccgcat caggcgctct tccgcttcct
     4081 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     4141 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     4201 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     4261 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     4321 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     4381 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     4441 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     4501 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     4561 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     4621 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     4681 acactagaag gacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     4741 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     4801 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     4861 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     4921 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     4981 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     5041 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     5101 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     5161 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     5221 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     5281 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     5341 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     5401 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     5461 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     5521 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     5581 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     5641 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     5701 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     5761 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     5821 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     5881 ttcaatatta ttgaagcatt tatcagggtt attgtctc
//
