LOCUS       sequence-232155-        5143 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  construction intermediate.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5143)
  AUTHORS   Peters JM, Koo BM, Patino R, Heussler GE, Hearne CC, Qu J, Inclan 
            YF, Hawkins JS, Lu CHS, Silvis MR, Harden MM, Osadnik H, Peters JE, 
            Engel JN, Dutton RJ, Grossman AD, Gross CA, Rosenberg OS
  TITLE     Enabling genetic analysis of diverse bacteria with Mobile-CRISPRi.
  JOURNAL   Nat Microbiol. 2019 Jan 7. pii: 10.1038/s41564-018-0327-z. doi: 
            10.1038/s41564-018-0327-z.
  PUBMED    30617347
REFERENCE   2  (bases 1 to 5143)
  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: "Nat 
            Microbiol. 2019 Jan 7. pii: 10.1038/s41564-018-0327-z. doi: 
            10.1038/s41564-018-0327-z."
COMMENT     Sequence Label: pJMP1055
FEATURES             Location/Qualifiers
     source          1..5143
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     oriT            8..116
                     /note="incP origin of transfer"
     rep_origin      385..773
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     CDS             complement(join(1074..1865,1866..1934))
                     /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     1697..1716
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1935..2039)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2107..2125
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2163..2185)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2285..2304
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2718..2734
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     2719..2735
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     terminator      2763..2857
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     terminator      2960..3046
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     protein_bind    3097..3144
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             complement(3212..3448)
                     /codon_start=1
                     /gene="dhfrIIb from plasmid R388"
                     /product="E. coli plasmid-associated dihydrofolate 
                     reductase"
                     /label=TpR
                     /note="confers resistance to trimethoprim"
                     /translation="MGQSSDEANAPVAGQFALPLSATFGLGDRVRKKSGAAWQGQVVGW
                     YCTKLTPEGYAVESESHPGSVQIYPVAALERVA"
     promoter        complement(join(3774..3779,3780..3796,3797..3802))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     protein_bind    3893..3940
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     promoter        complement(join(4336..4342,4343..4359,4360..4365))
                     /label=trc promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     mobile_element  4827..4992
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
ORIGIN
        1 agcttatcgg ccagcctcgc agagcaggat tcccgttgag caccgccagg tgcgaataag
       61 ggacagtgaa gaaggaacac ccgctcgcgg gtgggcctac ttcacctatc ctgcccggct
      121 gacgccgttg gatacaccaa ggaaagtcta cacgaaccct ttggcaaaat cctgtatatc
      181 gtgcgaaaaa ggatggatat accgaaaaaa tcgctataat gaccccgaag cagggttatg
      241 cagcggaaag tataccttaa ggaatcccct gataacgcag gaaagaacat gtgagcaaaa
      301 ggccagcaaa aggccaggaa ccgtaaaaag gccgcactcc ctgcctctgt catcacgata
      361 ctgtgatgcc atggctaatt cccatgtcag ccgttaagtg ttcctgtgtc actcaaaatt
      421 gctttgagag gctctaaggg cttctcagtg cgttacatcc ctggcttgtt gtccacaacc
      481 gttaaacctt aaaagcttta aaagccttat atattctttt ttttcttata aaacttaaaa
      541 ccttagaggc tatttaagtt gctgatttat attaatttta ttgttcaaac atgagagctt
      601 agtacgtgaa acatgagagc ttagtacgtt agccatgaga gcttagtacg ttagccatga
      661 gggtttagtt cgttaaacat gagagcttag tacgttaaac atgagagctt agtacgtgaa
      721 acatgagagc ttagtacgta ctatcaacag gttgaactgc tgatcttcag atcctctacg
      781 ccggacgcat cgtggccgga tcttgcggcc gcaaaaatta aaaatgaagt tttaaatcaa
      841 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
      901 caataactgc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
      961 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
     1021 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
     1081 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
     1141 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
     1201 gcaatgatac cgcgggaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca
     1261 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
     1321 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
     1381 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
     1441 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
     1501 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
     1561 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
     1621 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
     1681 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
     1741 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
     1801 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
     1861 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
     1921 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
     1981 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
     2041 acatttcccc gaaaagtgcc acctgacgtc taagaaacca ttattatcat gacattaacc
     2101 tataaaaata ggcgtatcac gaggcccttt cgtctcgcgc gtttcggtga cgacggtgaa
     2161 aacctctgac acatgcagct cccggagacg gtcacagctt gtctgtaagc ggatgccggg
     2221 agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg ctggcttaac
     2281 tatgcggcat cagagcagat tgtactgaga gtgcaccata tgcggtgtga aataccgcac
     2341 agatgcgtaa ggagaaaata ccgcatcagg cgccattcgc cattcaggct gcgcaactgt
     2401 tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctagagga ccagccgcgt
     2461 aacctggcaa aatcggttac ggttgagtaa taaatggatg ccctgcgtaa gcaggtgtgg
     2521 gcggacaata aagtcttaaa ctgaacaaaa tagatctaaa ctatgacaat aaagtcttaa
     2581 actagacaga atagttgtaa actgaaatca gtccagttat gctgtgaaaa agcatactgg
     2641 acttttgtta tggctaaagc aaactcttca ttttctgaag tgcaaattgc ccgtcgtatt
     2701 aaagaggggc gtggggttcg aggtcgacgg tatcgataag ctagcttaat tagctgagct
     2761 tggactcctg ttgatagatc cagtaatgac ctcagaactc catctggatt tgttcagaac
     2821 gctcggttgc cgccgggcgt tttttattgg tgagaatcca agctagactg cgatgagtgg
     2881 cagggcgggg cgtaattttt ttaaggcagt tattggtgcc cttaaacgcc tggggtaatg
     2941 actctctagc ttgaggcatc aaataaaacg aaaggctcag tcgaaagact gggcctttcg
     3001 ttttatctgt tgtttgtcgg tgaacgctct cctgagtagg acaaatccgc cgctaggagc
     3061 ttgcggcccg gacgatgagc tcgaattggg gatcttgaag tacctattcc gaagttccta
     3121 ttctctagaa agtataggaa cttcagagcg cttttgaagc tgatgctcga gtgcttaaaa
     3181 acttactcaa tggaataatt ctagataatt cttaggccac acgttcaagt gcagccacag
     3241 gataaatttg cactgagcct gggtgggatt cggactcgac cgcatagcct tcaggagtga
     3301 gttttgtgca ataccaaccg acgacttgac cctgccaagc ggcaccagat ttcttgcgta
     3361 cgcgatcccc taagccaaag gtggcactca ggggaagcgc aaactgccct gcaacgggag
     3421 cgttggcttc atcgctactt tgacccatgt cgaatccttc ttgtgaatct attatggcga
     3481 caagcaaatc gagctctgac tgcctacccc acaacaacta tcagaaagca ccagcacaac
     3541 ggctgcctaa ctttgtttta gggcgactgc cctgctgcgt aacatcgttg ctgctccata
     3601 acatcaaaca tcgacccacg gcgtaacgcg cttgctgctt ggatgcccga ggcatagact
     3661 gtacaaaaaa acagtcataa caagccatga aaaccgccac tgcgccgtta ccaccgctgc
     3721 gttcggtcaa ggttctggac cagttgcgtg agcgcatacg ctacttgcat tacagtttac
     3781 gaaccgaaca ggcttatgtc aactgggttc gtgaattatc cattgctgtt gacaaaggga
     3841 atcaggctcg agagttcaac agacagcctt cgttcttatg gccgttcagg aggaagttcc
     3901 tattccgaag ttcctattct ctagaaagta taggaacttc ttgagaagag aaaagaaaac
     3961 cgccgatcct gtccaccgca ttactgcaag gtagtggaca agaccggcgg tcttaagttt
     4021 tttggctgaa acgaatgacg aaaaggctgt ctgagcaaat ccaggaggtc gtttaaaccg
     4081 tattagcccg gtattttgga aatagcggaa gcactgactt ttgttatcaa taaaaaaggc
     4141 cccccgttag ggaggcctta ttgttcgtct actcggaaga gcgagagaca acagaacggt
     4201 cagccacatg aattcaaaaa aaaagcaccg actcggtgcc actttttcaa gttgataacg
     4261 gactagcctt atttaaactt gctatgctgt ttccagcata gctcttaaac agaccgctaa
     4321 actgaaagtt ccacacatta tacgagccgg atgattaatt gtcaacagct catttcagaa
     4381 tatttgccag aaccggaatt ctttcagctc agtcgatagg tagtaggcaa gagtagtcgc
     4441 acctttggtc gaaaaaaaaa gcccgcactg tcaggtgcgg gcttttttct gtgtttcccc
     4501 aaaagtaaaa acccgccgaa gcgggttttt acgtaaaaca ggtgaaactg accagacgag
     4561 aaggctttgg aggtgataat ggggctcaag gacccgggag gcagataggt aggcatggcc
     4621 cccattttca atacaagcaa cgcatgagaa agcccccgga agatcacctt ccgggggctt
     4681 ttttattgcg ctccggcaat taaaaaagcg gctaaccacg ccgctttttt tacgtctgca
     4741 gagctcatag gcaagcgaat cgtgatgcct cttagccagt actagtcgac atcggagaag
     4801 agtacggctc ttttaaccgc ctcaagaacc agataagtga aatctagttc caaactattt
     4861 tgtcattttt aattttcgta ttagcttacg acgctacacc cagttcccat ctattttgtc
     4921 actcttccct aaataatcct taaaaactcc atttccaccc ctcccagttc ccaactattt
     4981 tgtccgccca cagcggggca tttttcttcc tgttatgttt gggcgctgca ttaatgaatc
     5041 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     5101 gacccgctgc gctcggtcgt tcggctgcgg cgagcggtat cag
//
