LOCUS       sequence-305979-        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"
     primer_bind     86..105
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        86..104
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    105..129
                     /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      5584..5655
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     promoter        5669..5703
                     /label=BBa_J23108
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.51"
     terminator      5958..5985
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     promoter        6012..6116
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6117..6185,6186..6977)
                     /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(6335..6354)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7148..7736
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7637..7656
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(7946..9028)
                     /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     8990..9009
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(join(9029..9066,1..40))
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 atgccatacc gcgaaaggtt ttgcgccatt cgatggtgtc cgggatctcg acgctctccc
       61 ttatgcgact cctgcattag gaaattaata cgactcacta taggggaatt gtgagcggat
      121 aacaattccc ctgtagaaat aattttgttt aactttaata aggagatata ccatgaacag
      181 tcagcaaaat cctgatttag ctgttcatcc cttggcaatt cctatggaag gcttactagg
      241 agaaagtgct acaactcttg agaagaatgt aattgccaca caactctcag aggaagccca
      301 agtaaagcta gaggtaatcc aaagtttact ggaaccctgc gatcgcacaa cttatgggca
      361 aaagttgcgg gaagcagcag agaaactaaa tgtatcgttg cgaacggtac aaaggttggt
      421 gaaaaactgg gaacaagatg gcttagtcgg actcactcaa acaagtaggg ctgataaagg
      481 aaaacaccgc attggtgagt tttgggaaaa cttcattacc aaaacctaca aggagggtaa
      541 caagggaagt aaacgtatga cccctaaaca agttgctctc agagtcgagg ctaaagcccg
      601 tgaattaaaa gactctaagc cgcccaatta caaaaccgtg ttacgggtat tagcacccat
      661 tttggaaaag caacaaaaag ccaagagtat ccgcagtcct ggttggagag gaactacgct
      721 ttcggttaaa acccgtgaag gaaaagattt atcggttgat tacagtaacc atgtttggca
      781 atgtgaccat acccgcgtgg atgtgttgct ggtagatcaa catggtgaaa ttttaagtcg
      841 tccctggcta acaacagtaa ttgatactta ctctcgttgc attatgggta tcaacttggg
      901 ctttgatgca cccagttctg gggtagtagc attagcgtta cgccatgcaa ttctaccaaa
      961 gcgttacggt tccgagtaca aactgcattg tgagtgggga acctatggaa aaccagaaca
     1021 tttttatact gatggcggta aagactttcg ctctaaccac ttgagtcaga ttggggcgca
     1081 attgggattt gtctgtcatt tacgcgatcg cccttctgaa ggtggagtag tagaacgtcc
     1141 cttcaaaaca ttaaatgacc aactattttc aacgcttcct gggtacaccg gatctaatgt
     1201 gcaggaacgc ccagaagatg cagagaagga cgcaagactt actttgcgag aactagaaca
     1261 gttacttgtg cgttacatcg tagatcgtta caaccaaagt attgatgcgc ggatgggcga
     1321 ccaaacgcgc tttgagcgtt gggaagcagg attgcctaca gtgccagtac caataccaga
     1381 acgagatttg gatatttgtt taatgaagca gtcacggcgc actgtgcaaa gaggtggttg
     1441 tttgcagttt cagaatttaa tgtatcgggg ggaatatttg gcaggttatg ccggagaaac
     1501 tgtcaactta aggtttgacc ccagagacat tacaacaatt ttggtttatc gccaggaaaa
     1561 caatcaggaa gtatttctga ctcgcgctca cgctcaaggt ttggagacag agcaactggc
     1621 attagatgag gctgaggcag caagtcgcag actccgtacc gcagggaaaa ctatcagtaa
     1681 ccaatcatta ttgcaagaag ttgttgaccg cgatgctctt gtcgctacca agaaaagccg
     1741 taaggagcgt caaaaattgg aacagactgt tttgcgatct gctgctgttg atgaaagtaa
     1801 tagagaatcc ttgccttctc aaatagttga accagatgaa gtggaatcta cagaaacggt
     1861 tcactctcaa tacgaagaca ttgaggtgtg ggactatgaa caacttcgtg aagaatatgg
     1921 gttttaaaca atgacagaag ctcaggcgat cgccaagcag ttgggtgggg taaaaccgga
     1981 tgatgagtgg ttacaagctg aaattgctcg tctcaagggt aagagcattg tgcctttaca
     2041 gcaggtaaaa actctccatg attggttaga tggcaagcgc aaggcaagaa aatcttgccg
     2101 agtagttggg gaatcgagaa ctggcaagac agttgcttgt gatgcctaca gatacaggca
     2161 caaacctcag caggaagctg gacgacctcc aactgtgcct gtcgtttata ttcgacctca
     2221 ccaaaaatgt ggccccaagg atttgtttaa aaagattact gagtacctca agtatcgggt
     2281 aacaaaaggg actgtatctg attttcgaga taggacgata gaagtactca agggttgtgg
     2341 cgtagagatg ctaattattg atgaagctga ccgtctcaag cctgaaactt ttgctgatgt
     2401 gcgagatatt gccgaagatt taggaattgc tgtggtactg gtaggaacag accgtttgga
     2461 tgcggtaatt aagcgggatg agcaggttct cgaacgcttt cgggcgcatc ttcgctttgg
     2521 taaattgtcg ggagaggatt ttaagaacac cgtagaaatg tgggaacaaa tggttttgaa
     2581 actgccagta tcttctaatc taaagagcaa ggagatgcta cggattctca cgtcagcaac
     2641 tgaaggctac attggtcgcc ttgatgagat tcttagggaa gctgcaattc gttccttatc
     2701 aagaggattg aagaagattg acaaggctgt tttacaggaa gtagctaagg agtacaaatg
     2761 atagaagcac cagatgttaa accttggcta ttcttgatta aaccctatga aggggaaagc
     2821 ctgagccact ttcttggcag gttcagacgt gccaaccatt tatccgcaag tggattgggt
     2881 actttggcag gaattggtgc tatagtggca cgttgggaaa gatttcattt taatcctcgc
     2941 cctagtcagc aagaattgga agcgatcgca tctgtagtag aagtggatgc tcaaaggtta
     3001 gcccagatgt taccgcctgc tggagtggga atgcagcatg agccaattcg cttgtgtggg
     3061 gcttgttatg ccgagtcgcc ttgtcaccga attgaatggc agtacaagtc ggtgtggaag
     3121 tgcgatcgcc atcaactcaa gattttagca aagtgtccaa actgtcaagc accttttaaa
     3181 atgcctgcgc tgtgggagga tgggtgctgt cacagatgta ggatgccgtt tgcagaaatg
     3241 gcaaagctac agaaggtttg atgataaaac cagaaaaagg tgtgaaatta actaagtccc
     3301 tgaattgatc tggttgtcca aaaaatttgt gcgatcgcat ggcaagatta ttcctactat
     3361 tgatgtggcg tgagtcggat aacttgctct aatgggatcc cggttaatac caatagatca
     3421 gatatgagta ataaacatta cgacctttac ctgtggtatt cactgttggc acgacaacac
     3481 ctttatcgcg ccaatactgt agctgacgac gcgaacaatt agtgatttga gccgcttctg
     3541 tgcttgtgta gaattttcct tccatagaaa tcatcctaca gaagtagttt tttaaattta
     3601 cattttttaa aagcacatta tgtattcatg tgatattatt tttttatgag tcaaataact
     3661 attcaagctc gacttatttc ctttgaatca aaccgccaac aactctggaa gttgatggca
     3721 gatttaaaca cgccgttaat taacgaactg ctttgccagt taggtcaaca ccccgacttc
     3781 gagaagtggc aacaaaaggg taaactcccg tctaccgttg tgagccagtt atgtcaacct
     3841 ctcaaaactg accctcgctt tgcaggtcag cccagccgtt tatatatgtc ggcaattcat
     3901 attgtggact acatctacaa gtcctggctg gctatacaga aacggcttca acagcagcta
     3961 gatggaaaga cgcgctggct agaaatgctc aatagcgatg ctgaattagt agaacttagt
     4021 ggtgacactt tagaggctat tcgtgtcaaa gctgctgaaa ttttggcaat agctatgcca
     4081 gcatctgagt cagatagcgc ttcacctaaa gggaaaaaag gtaaaaagga gaaaaaaccc
     4141 tcatcttcta gccctaagcg tagtttatcc aagacattat ttgacgctta ccaagaaacg
     4201 gaagatatca agagccgtag cgccatcagc tacctgttaa aaaatggctg caaacttact
     4261 gacaaagaag aagattcaga aaaatttgct aaacgtcgtc gtcaagttga aatccaaatt
     4321 caaaggctta ccgaaaagtt aataagtcgg atgcctaaag gtcgagattt gaccaatgct
     4381 aaatggttgg agacactctt gactgctaca accactgttg ctgaagacaa cgcccaagcc
     4441 aaacgctggc aggatattct gttaactcga tcaagttctc tcccattccc ccttgttttt
     4501 gaaaccaacg aggatatggt ttggtcaaag aatcaaaagg gtaggctgtg tgttcacttc
     4561 aatggcttaa gcgatttaat ttttgaggtg tactgcggca atcgtcaact tcactggttt
     4621 caacgcttcc tagaagacca acagactaaa cgcaaaagca aaaatcagca ttctagcggc
     4681 ttgttcacac tcagaaatgg tcatctagtt tggcttgaag gtgagggtaa aggggaacct
     4741 tggaatcttc accacttgac cctttactgc tgtgttgaca atcgcttgtg gacagaggag
     4801 ggaacagaaa tcgttcgcca agagaaagca gatgaaatta ctaaattcat cacaaacatg
     4861 aagaagaaaa gcgatctaag cgatacacag caagctttga ttcaacgtaa acaatcaaca
     4921 cttactcgaa taaacaattc ctttgagcgt cctagccaac ccctttatca aggtcaatca
     4981 cacattttgg ttggagtaag cctgggacta gaaaaacctg ccacagtagc agtagtagat
     5041 gcgatcgcca acaaagtctt ggcttaccgg agtattaaac aattacttgg cgacaattac
     5101 gaactgctaa atcgccagag acgacaacag cagtacctat ctcacgaacg ccacaaagca
     5161 caaaaaaact tctctcccaa tcaatttgga gcatctgagt tagggcaaca tatagacaga
     5221 ttattagcta aagcaattgt agcgttagcg agaacctaca aagctggcag tattgtcttg
     5281 cccaagttag gggatatgcg ggaggttgtc caaagtgaaa ttcaagctat agcagaacaa
     5341 aaatttcccg gttatattga aggtcagcaa aaatatgcca aacagtaccg ggttaatgtt
     5401 catcggtgga gctacggcag attaattcaa agcattcaaa gtaaagcagc tcaaacagga
     5461 attgtgattg aggagggaaa acaacctatt cgaggtagtc cccacgacaa agcaaaggaa
     5521 ttagcacttt ctgcttacaa tctccgccta actaggcgaa gttaagtaag gatctccagg
     5581 catcaaataa aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg
     5641 tcggtgaacg ctctctacta gagagaccct gacagctagc tcagtcctag gtataatgct
     5701 agcatattaa tagcgccgca attcatgctg cttgcagcct ctgaattttg ttaaatgagg
     5761 gttagtttga ctgtataaat acagtcttgc tttctgaccc tggtagctgc tcaccctgat
     5821 gctgctgtca atagacagga taggtgcgct cccagcaata agggcgcgga tgtactgctg
     5881 tagtggctac tgaatcaccc ccgatcaagg gggaacccta aatgggttga aagttttatg
     5941 ttcagataat gcccgatggc tcaccttcgg gtgggccttt ctgcgcaggt ggcacttttc
     6001 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     6061 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
     6121 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     6181 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
     6241 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     6301 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta
     6361 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     6421 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     6481 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
     6541 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     6601 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     6661 tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     6721 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     6781 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     6841 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     6901 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     6961 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa
     7021 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     7081 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     7141 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     7201 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     7261 ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc
     7321 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     7381 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     7441 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     7501 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     7561 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     7621 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     7681 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     7741 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct caacgccaag
     7801 ctccaaaacg atctcaagaa gatcatctta ttaattcatg ccccgcgccc accggaagga
     7861 gctgactggg ttgaaggctc tcaagggcat cggtcgagat cccggtgcct aatgagtgag
     7921 ctaacttaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
     7981 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgcca
     8041 gggtggtttt tcttttcacc agtgagacgg gcaacagctg attgcccttc accgcctggc
     8101 cctgagagag ttgcagcaag cggtccacgc tggtttgccc cagcaggcga aaatcctgtt
     8161 tgatggtggt taacggcggg atataacatg agctgtcttc ggtatcgtcg tatcccacta
     8221 ccgagatgtc cgcaccaacg cgcagcccgg actcggtaat ggcgcgcatt gcgcccagcg
     8281 ccatctgatc gttggcaacc agcatcgcag tgggaacgat gccctcattc agcatttgca
     8341 tggtttgttg aaaaccggac atggcactcc agtcgccttc ccgttccgct atcggctgaa
     8401 tttgattgcg agtgagatat ttatgccagc cagccagacg cagacgcgcc gagacagaac
     8461 ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa tgcgaccaga tgctccacgc
     8521 ccagtcgcgt accgtcttca tgggagaaaa taatactgtt gatgggtgtc tggtcagaga
     8581 catcaagaaa taacgccgga acattagtgc aggcagcttc cacagcaatg gcatcctggt
     8641 catccagcgg atagttaatg atcagcccac tgacgcgttg cgcgagaaga ttgtgcaccg
     8701 ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga caccaccacg ctggcaccca
     8761 gttgatcggc gcgagattta atcgccgcga caatttgcga cggcgcgtgc agggccagac
     8821 tggaggtggc aacgccaatc agcaacgact gtttgcccgc cagttgttgt gccacgcggt
     8881 tgggaatgta attcagctcc gccatcgccg cttccacttt ttcccgcgtt ttcgcagaaa
     8941 cgtggctggc ctggttcacc acgcgggaaa cggtctgata agagacaccg gcatactctg
     9001 cgacatcgta taacgttact ggtttcacat tcaccaccct gaattgactc tcttccgggc
     9061 gctatc
//
