LOCUS       sequence-342310-        4181 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial genomic integration vector, CymR repressor.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4181)
  AUTHORS   Hao N, Chen Q, Dodd IB, Shearwin KE
  TITLE     The pIT5 Plasmid Series, an Improved Toolkit for Repeated Genome 
            Integration in E. coli.
  JOURNAL   ACS Synth Biol. 2021 Jun 30. doi: 10.1021/acssynbio.1c00215.
  PUBMED    34190535
REFERENCE   2  (bases 1 to 4181)
  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. 2021 Jun 30. doi: 10.1021/acssynbio.1c00215."
COMMENT     Sequence Label: pIT5_KW_CymR
FEATURES             Location/Qualifiers
     source          1..4181
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    27..54
                     /label=CuO
                     /bound_moiety="CymR"
                     /note="CymR-binding P2 operator sequence from the p-cmt 
                     operon of Pseudomonas putida (Mullick et al., 2006)"
     protein_bind    85..112
                     /label=CuO
                     /bound_moiety="CymR"
                     /note="CymR-binding P2 operator sequence from the p-cmt 
                     operon of Pseudomonas putida (Mullick et al., 2006)"
     misc_RNA        119..168
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     terminator      1037..1064
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(1113..1144)
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     protein_bind    1220..1253
                     /label=lox66
                     /bound_moiety="Cre recombinase"
                     /note="Right element (RE) mutant of loxP (Araki et al., 
                     2010). Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(1417..1436)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1428..2243
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(1501..1520)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2365..2753
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     protein_bind    2768..2801
                     /label=lox71
                     /bound_moiety="Cre recombinase"
                     /note="Left element (LE) mutant of loxP (Araki et al., 
                     2010). Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     terminator      complement(3070..3316)
                     /label=lambda tL3 terminator
                     /note="transcription terminator tL3 from phage lambda"
     primer_bind     complement(3407..3429)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3580..3598
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      3639..3725
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3822..3908
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      4005..4091
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 gaattccgct taacgatcgt tggctgaaca aacagacaat ctggtctgtt tgtattatgg
       61 aaaatttttc tgtataatag attcaacaaa cagacaatct ggtctgtttg tattatagct
      121 gtcaccggat gtgctttccg gtctgatgag tccgtgagga cgaaacagcc tctacaaata
      181 attttgttta atctagactc gagactagtc tcggtaccaa attccagaaa agaggcctcc
      241 cgaaaggggg gccttttttc gttttggtcc aatggcggcg cgccatcgaa tggtgcaaaa
      301 cctttcgcgg tatggcatga tagcgcccgg aagagagtca attcagggtg gtgaatatga
      361 gcccgaaacg tcgtacccag gcagaacgtg caatggaaac ccagggtaaa ctgattgcag
      421 cagcactggg tgttctgcgt gaaaaaggtt atgcaggttt tcgtattgca gatgttccgg
      481 gtgcagccgg tgttagccgt ggtgcacaga gccatcattt tccgaccaaa ctggaactgc
      541 tgctggcaac ctttgaatgg ctgtatgagc agattaccga acgtagccgt gcacgtctgg
      601 caaaactgaa accggaagat gatgttattc agcagatgct ggatgatgca gcagaatttt
      661 ttctggatga tgattttagc atcggcctgg atctgattgt tgcagcagat cgtgatccgg
      721 cactgcgtga aggtattcag cgtaccgttg aacgtaatcg ttttgttgtt gaagatatgt
      781 ggctgggtgt gctggtgagc cgtggtctga gccgtgatga tgccgaagat attctgtggc
      841 tgatttttaa cagcgttcgt ggtctggtag ttcgtagcct gtggcagaaa gataaagaac
      901 gttttgaacg tgtgcgtaat agcaccctgg aaattgcacg tgaacgttat gcaaaattca
      961 aacgttgata aggatcctaa ttggtaacga atcaatgcat ggcgcctaac ctaaactgac
     1021 aggcatcaaa ttaagcagaa ggccatcctg acggatggcc tttttgcgtt tcgaacaatt
     1081 gaaaaaacct cgcgccttac ctgttgagta atagtcaaaa gcctccggtc ggaggctttt
     1141 gactttctgc ttactgaatt tcggtggtgc cgttaattaa ccggtgggcc ctcatgataa
     1201 taatggtttc ttagacgtct accgttcgta taatgtatgc tatacgaagt tatcctaggc
     1261 gacaatgtaa ctactagagc tgatccttca actcagcaaa agttcgattt attcaacaaa
     1321 gccacgttgt gtctcaaaat ctctgatgtt acattgcaca agataaaaat atatcatcat
     1381 gaacaataaa actgtctgct tacataaaca gtaatacaag gggtgttatg agccatattc
     1441 aacgggaaac gtcttgctcc cgtccgcgct taaactccaa catggacgct gatttatatg
     1501 ggtataaatg ggctcgcgat aatgtcgggc aatcaggtgc gacaatctat cgcttgtatg
     1561 ggaagcccga tgcgccagag ttgtttctga aacatggcaa aggtagcgtt gccaatgatg
     1621 ttacagatga gatggtccgt ctcaactggc tgacggagtt tatgcctctc ccgaccatca
     1681 agcattttat ccgtactcct gatgatgcgt ggttactcac caccgcgatt cctgggaaaa
     1741 cagccttcca ggtattagaa gaatatcctg attcaggtga aaatattgtt gatgcgctgg
     1801 ccgtgttcct gcgccggtta cattcgattc ctgtttgtaa ttgtcctttt aacagcgatc
     1861 gtgtatttcg tcttgctcag gcgcaatcac gcatgaataa cggtttggtt gatgcgagtg
     1921 attttgatga cgagcgtaat ggctggcctg ttgaacaagt ctggaaagaa atgcacaagc
     1981 tcttgccatt ctcaccggat tcagtcgtca ctcatggtga tttctcactt gataacctta
     2041 tttttgacga ggggaaatta ataggttgta ttgatgttgg acgggtcgga atcgcagacc
     2101 gttaccagga ccttgccatt ctttggaact gcctcggtga gttttctcct tcattacaga
     2161 aacggctttt tcaaaaatat ggtattgata atcctgatat gaataaattg cagtttcatt
     2221 tgatgctcga tgagtttttc taataatact agcagaaatc atccttagcg aaagctaagg
     2281 atttttttta tctgattacc gcctttgagt gagcgtcgac ctagtgcggc cgcaagatcc
     2341 ggccacgatg cgtccggcgt agaggatctg aagatcagca gttcaacctg ttgatagtac
     2401 gtactaagct ctcatgtttc acgtactaag ctctcatgtt taacgtacta agctctcatg
     2461 tttaacgaac taaaccctca tggctaacgt actaagctct catggctaac gtactaagct
     2521 ctcatgtttc acgtactaag ctctcatgtt tgaacaataa aattaatata aatcagcaac
     2581 ttaaatagcc tctaaggttt taagttttat aagaaaaaaa agaatatata aggcttttaa
     2641 agcttttaag gtttaacggt tgtggacaac aagccaggga tgtaacgcac tgagaagccc
     2701 ttagagcctc tcaaagcaat tttgagtgac acaggaacac ttaacggctg acatgggaat
     2761 taggaagata acttcgtata atgtatgcta tacgaacggt acatatgcca tggtggtcga
     2821 tgtggacgct atgtggacag tggttgatat aaatccattt atatcatcag gttaggtgct
     2881 tttttgtgac accatccctg tcttccccca catgatgtgg gggttttttt tatcaatcac
     2941 ttacatcggt ttccgtaaga aaacagtaac ttaagaaaac ctataaggga ctggaagggc
     3001 tacaaagttc tgagagtgtg gacactatgt ggacactcag aagggaaatg ctagcttctt
     3061 cgtctgtttc tactggtatt ggcacaaacc tgattccaat ttgagcaagg ctatgtgcca
     3121 tctcgatact cgttcttaac tcaacagaag atgctttgtg catacagccc ctcgtttatt
     3181 atttatctcc tcagccagcc gctgtgcttt cagtggattt cggataacag aaaggccggg
     3241 aaatacccag cctcgctttg taacggagta gagacgaaag tgattgcgcc tacccggata
     3301 ttatcgtgag gatgcgtcat cgccattaat tcactgatca gtgataagct gtcaaacatg
     3361 agaattgatc cggctgcctc gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc
     3421 agctcccgga gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc
     3481 agggcgcgtc agcgggtgtt ggcgggtgtc ggggcgcagc catgacccag tcacgtagcg
     3541 atagcggagt gtatgctgca catgacatta acctataaaa ataggcgtat cacgaggccc
     3601 tttcgtcttc aagaattaat tcccaattcc ccaggcatca aataaaacga aaggctcagt
     3661 cgaaagactg ggcctttcgt tttatctgtt gtttgtcggt gaacgctctc ctgagtagga
     3721 caaatccgcc gggagcggat ttgaacgttg cgaagcaacg gcccggaggg tggcgggcag
     3781 gacgcccgcc ataaactgcc aggaattaat tccccaggca tcaaataaaa cgaaaggctc
     3841 agtcgaaaga ctgggccttt cgttttatct gttgtttgtc ggtgaacgct ctcctgagta
     3901 ggacaaatcc gccgggagcg gatttgaacg ttgcgaagca acggcccgga gggtggcggg
     3961 caggacgccc gccataaact gccaggaatt aattccccag gcatcaaata aaacgaaagg
     4021 ctcagtcgaa agactgggcc tttcgtttta tctgttgttt gtcggtgaac gctctcctga
     4081 gtaggacaaa tccgccggga gcggatttga acgttgcgaa gcaacggccc ggagggtggc
     4141 gggcaggacg cccgccataa actgccagga attggggatc g
//
