LOCUS       sequence-272867-        9066 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses ShCAST under the T7 promoter and an empty sgRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9066)
  AUTHORS   Zhang Y, Sun X, Wang Q, Xu J, Dong F, Yang S, Yang J, Zhang Z, Qian 
            Y, Chen J, Zhang J, Liu Y, Tao R, Jiang Y, Yang J, Yang S
  TITLE     Multicopy chromosomal integration using CRISPR-associated 
            transposases.
  JOURNAL   ACS Synth Biol. 2020 Jun 17. doi: 10.1021/acssynbio.0c00073.
  PUBMED    32551502
REFERENCE   2  (bases 1 to 9066)
  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: "ACS Synth 
            Biol. 2020 Jun 17. doi: 10.1021/acssynbio.0c00073."
COMMENT     Sequence Label: pHelper-T7IS1
FEATURES             Location/Qualifiers
     source          1..9066
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(111..1193)
                     /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     1155..1174
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1194..1271)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1317..1336
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1317..1335
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1336..1360
                     /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)."
     terminator      6815..6886
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     promoter        6900..6934
                     /label=BBa_J23108
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.51"
     terminator      7189..7216
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     promoter        7243..7347
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7348..7416,7417..8208)
                     /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     complement(7566..7585)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8379..8967
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8868..8887
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tcatgccccg cgcccaccgg aaggagctga ctgggttgaa ggctctcaag ggcatcggtc
       61 gagatcccgg tgcctaatga gtgagctaac ttacattaat tgcgttgcgc tcactgcccg
      121 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
      181 gaggcggttt gcgtattggg cgccagggtg gtttttcttt tcaccagtga gacgggcaac
      241 agctgattgc ccttcaccgc ctggccctga gagagttgca gcaagcggtc cacgctggtt
      301 tgccccagca ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata acatgagctg
      361 tcttcggtat cgtcgtatcc cactaccgag atgtccgcac caacgcgcag cccggactcg
      421 gtaatggcgc gcattgcgcc cagcgccatc tgatcgttgg caaccagcat cgcagtggga
      481 acgatgccct cattcagcat ttgcatggtt tgttgaaaac cggacatggc actccagtcg
      541 ccttcccgtt ccgctatcgg ctgaatttga ttgcgagtga gatatttatg ccagccagcc
      601 agacgcagac gcgccgagac agaacttaat gggcccgcta acagcgcgat ttgctggtga
      661 cccaatgcga ccagatgctc cacgcccagt cgcgtaccgt cttcatggga gaaaataata
      721 ctgttgatgg gtgtctggtc agagacatca agaaataacg ccggaacatt agtgcaggca
      781 gcttccacag caatggcatc ctggtcatcc agcggatagt taatgatcag cccactgacg
      841 cgttgcgcga gaagattgtg caccgccgct ttacaggctt cgacgccgct tcgttctacc
      901 atcgacacca ccacgctggc acccagttga tcggcgcgag atttaatcgc cgcgacaatt
      961 tgcgacggcg cgtgcagggc cagactggag gtggcaacgc caatcagcaa cgactgtttg
     1021 cccgccagtt gttgtgccac gcggttggga atgtaattca gctccgccat cgccgcttcc
     1081 actttttccc gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg ggaaacggtc
     1141 tgataagaga caccggcata ctctgcgaca tcgtataacg ttactggttt cacattcacc
     1201 accctgaatt gactctcttc cgggcgctat catgccatac cgcgaaaggt tttgcgccat
     1261 tcgatggtgt ccgggatctc gacgctctcc cttatgcgac tcctgcatta ggaaattaat
     1321 acgactcact ataggggaat tgtgagcgga taacaattcc cctgtagaaa taattttgtt
     1381 taactttaat aaggagatat accatgaaca gtcagcaaaa tcctgattta gctgttcatc
     1441 ccttggcaat tcctatggaa ggcttactag gagaaagtgc tacaactctt gagaagaatg
     1501 taattgccac acaactctca gaggaagccc aagtaaagct agaggtaatc caaagtttac
     1561 tggaaccctg cgatcgcaca acttatgggc aaaagttgcg ggaagcagca gagaaactaa
     1621 atgtatcgtt gcgaacggta caaaggttgg tgaaaaactg ggaacaagat ggcttagtcg
     1681 gactcactca aacaagtagg gctgataaag gaaaacaccg cattggtgag ttttgggaaa
     1741 acttcattac caaaacctac aaggagggta acaagggaag taaacgtatg acccctaaac
     1801 aagttgctct cagagtcgag gctaaagccc gtgaattaaa agactctaag ccgcccaatt
     1861 acaaaaccgt gttacgggta ttagcaccca ttttggaaaa gcaacaaaaa gccaagagta
     1921 tccgcagtcc tggttggaga ggaactacgc tttcggttaa aacccgtgaa ggaaaagatt
     1981 tatcggttga ttacagtaac catgtttggc aatgtgacca tacccgcgtg gatgtgttgc
     2041 tggtagatca acatggtgaa attttaagtc gtccctggct aacaacagta attgatactt
     2101 actctcgttg cattatgggt atcaacttgg gctttgatgc acccagttct ggggtagtag
     2161 cattagcgtt acgccatgca attctaccaa agcgttacgg ttccgagtac aaactgcatt
     2221 gtgagtgggg aacctatgga aaaccagaac atttttatac tgatggcggt aaagactttc
     2281 gctctaacca cttgagtcag attggggcgc aattgggatt tgtctgtcat ttacgcgatc
     2341 gcccttctga aggtggagta gtagaacgtc ccttcaaaac attaaatgac caactatttt
     2401 caacgcttcc tgggtacacc ggatctaatg tgcaggaacg cccagaagat gcagagaagg
     2461 acgcaagact tactttgcga gaactagaac agttacttgt gcgttacatc gtagatcgtt
     2521 acaaccaaag tattgatgcg cggatgggcg accaaacgcg ctttgagcgt tgggaagcag
     2581 gattgcctac agtgccagta ccaataccag aacgagattt ggatatttgt ttaatgaagc
     2641 agtcacggcg cactgtgcaa agaggtggtt gtttgcagtt tcagaattta atgtatcggg
     2701 gggaatattt ggcaggttat gccggagaaa ctgtcaactt aaggtttgac cccagagaca
     2761 ttacaacaat tttggtttat cgccaggaaa acaatcagga agtatttctg actcgcgctc
     2821 acgctcaagg tttggagaca gagcaactgg cattagatga ggctgaggca gcaagtcgca
     2881 gactccgtac cgcagggaaa actatcagta accaatcatt attgcaagaa gttgttgacc
     2941 gcgatgctct tgtcgctacc aagaaaagcc gtaaggagcg tcaaaaattg gaacagactg
     3001 ttttgcgatc tgctgctgtt gatgaaagta atagagaatc cttgccttct caaatagttg
     3061 aaccagatga agtggaatct acagaaacgg ttcactctca atacgaagac attgaggtgt
     3121 gggactatga acaacttcgt gaagaatatg ggttttaaac aatgacagaa gctcaggcga
     3181 tcgccaagca gttgggtggg gtaaaaccgg atgatgagtg gttacaagct gaaattgctc
     3241 gtctcaaggg taagagcatt gtgcctttac agcaggtaaa aactctccat gattggttag
     3301 atggcaagcg caaggcaaga aaatcttgcc gagtagttgg ggaatcgaga actggcaaga
     3361 cagttgcttg tgatgcctac agatacaggc acaaacctca gcaggaagct ggacgacctc
     3421 caactgtgcc tgtcgtttat attcgacctc accaaaaatg tggccccaag gatttgttta
     3481 aaaagattac tgagtacctc aagtatcggg taacaaaagg gactgtatct gattttcgag
     3541 ataggacgat agaagtactc aagggttgtg gcgtagagat gctaattatt gatgaagctg
     3601 accgtctcaa gcctgaaact tttgctgatg tgcgagatat tgccgaagat ttaggaattg
     3661 ctgtggtact ggtaggaaca gaccgtttgg atgcggtaat taagcgggat gagcaggttc
     3721 tcgaacgctt tcgggcgcat cttcgctttg gtaaattgtc gggagaggat tttaagaaca
     3781 ccgtagaaat gtgggaacaa atggttttga aactgccagt atcttctaat ctaaagagca
     3841 aggagatgct acggattctc acgtcagcaa ctgaaggcta cattggtcgc cttgatgaga
     3901 ttcttaggga agctgcaatt cgttccttat caagaggatt gaagaagatt gacaaggctg
     3961 ttttacagga agtagctaag gagtacaaat gatagaagca ccagatgtta aaccttggct
     4021 attcttgatt aaaccctatg aaggggaaag cctgagccac tttcttggca ggttcagacg
     4081 tgccaaccat ttatccgcaa gtggattggg tactttggca ggaattggtg ctatagtggc
     4141 acgttgggaa agatttcatt ttaatcctcg ccctagtcag caagaattgg aagcgatcgc
     4201 atctgtagta gaagtggatg ctcaaaggtt agcccagatg ttaccgcctg ctggagtggg
     4261 aatgcagcat gagccaattc gcttgtgtgg ggcttgttat gccgagtcgc cttgtcaccg
     4321 aattgaatgg cagtacaagt cggtgtggaa gtgcgatcgc catcaactca agattttagc
     4381 aaagtgtcca aactgtcaag caccttttaa aatgcctgcg ctgtgggagg atgggtgctg
     4441 tcacagatgt aggatgccgt ttgcagaaat ggcaaagcta cagaaggttt gatgataaaa
     4501 ccagaaaaag gtgtgaaatt aactaagtcc ctgaattgat ctggttgtcc aaaaaatttg
     4561 tgcgatcgca tggcaagatt attcctacta ttgatgtggc gtgagtcgga taacttgctc
     4621 taatgggatc ccggttaata ccaatagatc agatatgagt aataaacatt acgaccttta
     4681 cctgtggtat tcactgttgg cacgacaaca cctttatcgc gccaatactg tagctgacga
     4741 cgcgaacaat tagtgatttg agccgcttct gtgcttgtgt agaattttcc ttccatagaa
     4801 atcatcctac agaagtagtt ttttaaattt acatttttta aaagcacatt atgtattcat
     4861 gtgatattat ttttttatga gtcaaataac tattcaagct cgacttattt cctttgaatc
     4921 aaaccgccaa caactctgga agttgatggc agatttaaac acgccgttaa ttaacgaact
     4981 gctttgccag ttaggtcaac accccgactt cgagaagtgg caacaaaagg gtaaactccc
     5041 gtctaccgtt gtgagccagt tatgtcaacc tctcaaaact gaccctcgct ttgcaggtca
     5101 gcccagccgt ttatatatgt cggcaattca tattgtggac tacatctaca agtcctggct
     5161 ggctatacag aaacggcttc aacagcagct agatggaaag acgcgctggc tagaaatgct
     5221 caatagcgat gctgaattag tagaacttag tggtgacact ttagaggcta ttcgtgtcaa
     5281 agctgctgaa attttggcaa tagctatgcc agcatctgag tcagatagcg cttcacctaa
     5341 agggaaaaaa ggtaaaaagg agaaaaaacc ctcatcttct agccctaagc gtagtttatc
     5401 caagacatta tttgacgctt accaagaaac ggaagatatc aagagccgta gcgccatcag
     5461 ctacctgtta aaaaatggct gcaaacttac tgacaaagaa gaagattcag aaaaatttgc
     5521 taaacgtcgt cgtcaagttg aaatccaaat tcaaaggctt accgaaaagt taataagtcg
     5581 gatgcctaaa ggtcgagatt tgaccaatgc taaatggttg gagacactct tgactgctac
     5641 aaccactgtt gctgaagaca acgcccaagc caaacgctgg caggatattc tgttaactcg
     5701 atcaagttct ctcccattcc cccttgtttt tgaaaccaac gaggatatgg tttggtcaaa
     5761 gaatcaaaag ggtaggctgt gtgttcactt caatggctta agcgatttaa tttttgaggt
     5821 gtactgcggc aatcgtcaac ttcactggtt tcaacgcttc ctagaagacc aacagactaa
     5881 acgcaaaagc aaaaatcagc attctagcgg cttgttcaca ctcagaaatg gtcatctagt
     5941 ttggcttgaa ggtgagggta aaggggaacc ttggaatctt caccacttga ccctttactg
     6001 ctgtgttgac aatcgcttgt ggacagagga gggaacagaa atcgttcgcc aagagaaagc
     6061 agatgaaatt actaaattca tcacaaacat gaagaagaaa agcgatctaa gcgatacaca
     6121 gcaagctttg attcaacgta aacaatcaac acttactcga ataaacaatt cctttgagcg
     6181 tcctagccaa cccctttatc aaggtcaatc acacattttg gttggagtaa gcctgggact
     6241 agaaaaacct gccacagtag cagtagtaga tgcgatcgcc aacaaagtct tggcttaccg
     6301 gagtattaaa caattacttg gcgacaatta cgaactgcta aatcgccaga gacgacaaca
     6361 gcagtaccta tctcacgaac gccacaaagc acaaaaaaac ttctctccca atcaatttgg
     6421 agcatctgag ttagggcaac atatagacag attattagct aaagcaattg tagcgttagc
     6481 gagaacctac aaagctggca gtattgtctt gcccaagtta ggggatatgc gggaggttgt
     6541 ccaaagtgaa attcaagcta tagcagaaca aaaatttccc ggttatattg aaggtcagca
     6601 aaaatatgcc aaacagtacc gggttaatgt tcatcggtgg agctacggca gattaattca
     6661 aagcattcaa agtaaagcag ctcaaacagg aattgtgatt gaggagggaa aacaacctat
     6721 tcgaggtagt ccccacgaca aagcaaagga attagcactt tctgcttaca atctccgcct
     6781 aactaggcga agttaagtaa ggatctccag gcatcaaata aaacgaaagg ctcagtcgaa
     6841 agactgggcc tttcgtttta tctgttgttt gtcggtgaac gctctctact agagagaccc
     6901 tgacagctag ctcagtccta ggtataatgc tagcatatta atagcgccgc aattcatgct
     6961 gcttgcagcc tctgaatttt gttaaatgag ggttagtttg actgtataaa tacagtcttg
     7021 ctttctgacc ctggtagctg ctcaccctga tgctgctgtc aatagacagg ataggtgcgc
     7081 tcccagcaat aagggcgcgg atgtactgct gtagtggcta ctgaatcacc cccgatcaag
     7141 ggggaaccct aaatgggttg aaagttttat gttcagataa tgcccgatgg ctcaccttcg
     7201 ggtgggcctt tctgcgcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     7261 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     7321 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     7381 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     7441 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
     7501 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
     7561 gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc
     7621 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
     7681 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
     7741 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
     7801 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
     7861 ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta
     7921 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
     7981 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
     8041 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
     8101 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     8161 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
     8221 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     8281 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     8341 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     8401 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     8461 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     8521 actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     8581 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     8641 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     8701 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     8761 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     8821 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     8881 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     8941 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     9001 gccttttgct ggccttttgc tcaacgccaa gctccaaaac gatctcaaga agatcatctt
     9061 attaat
//
