LOCUS       sequence-232153-        5641 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 5641)
  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 5641)
  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: pJMP1054
FEATURES             Location/Qualifiers
     source          1..5641
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(7..885)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     promoter        912..1196
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     1119..1138
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     RBS             1223..1245
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     misc_feature    1271..1327
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     mobile_element  1368..1533
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     oriT            1692..1800
                     /note="incP origin of transfer"
     rep_origin      2069..2457
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     CDS             complement(join(2758..3549,3550..3618))
                     /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     3381..3400
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3619..3723)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3791..3809
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3847..3869)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3969..3988
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4402..4418
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     4403..4419
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     terminator      4447..4541
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     terminator      4644..4730
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     protein_bind    4781..4828
                     /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(4896..5132)
                     /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(5458..5463,5464..5480,5481..5486))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     protein_bind    5538..5585
                     /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)."
ORIGIN
        1 cccaaattat gacaacttga cggctacatc attcactttt tcttcacaac cggcacggaa
       61 ctcgctcggg ctggccccgg tgcatttttt aaatacccgc gagaaataga gttgatcgtc
      121 aaaaccaaca ttgcgaccga cggtggcgat aggcatccgg gtggtgctca aaagcagctt
      181 cgcctggctg atacgttggt cctcgcgcca gcttaagacg ctaatcccta actgctggcg
      241 gaaaagatgt gacagacgcg acggcgacaa gcaaacatgc tgtgcgacgc tggcgatatc
      301 aaaattgctg tctgccaggt gatcgctgat gtactgacaa gcctcgcgta cccgattatc
      361 catcggtgga tggagcgact cgttaatcgc ttccatgcgc cgcagtaaca attgctcaag
      421 cagatttatc gccagcagct ccgaatagcg cccttcccct tgcccggcgt taatgatttg
      481 cccaaacagg tcgctgaaat gcggctggtg cgcttcatcc gggcgaaaga accccgtatt
      541 ggcaaatatt gacggccagt taagccattc atgccagtag gcgcgcggac gaaagtaaac
      601 ccactggtga taccattcgc gagcctccgg atgacgaccg tagtgatgaa tctctcctgg
      661 cgggaacagc aaaatatcac ccggtcggca aacaaattct cgtccctgat ttttcaccac
      721 cccctgaccg cgaatggtga gattgagaat ataacctttc attcccagcg gtcggtcgat
      781 aaaaaaatcg agataaccgt tggcctcaat cggcgttaaa cccgccacca gatgggcatt
      841 aaacgagtat cccggcagca ggggatcatt ttgcgcttca gccatacttt tcatactccc
      901 gccattcaga gaagaaacca attgtccata ttgcatcaga cattgccgtc actgcgtctt
      961 ttactggctc ttctcgctaa ccaaaccggt aaccccgctt attaaaagca ttctgtaaca
     1021 aagcgggacc aaagccatga caaaaacgcg taacaaaagt gtctataatc acggcagaaa
     1081 agtccacatt gattatttgc acggcgtcac actttgctat gccatagcat ttttatccat
     1141 aagattagcg gatcctacct gacgcttttt atcgcaactc tctactgttt ctccataccc
     1201 gtttttttgg gctagaaata attttgttta actttaagaa ggagatatac atacccatgg
     1261 gatctgataa gaattcgagc tcggtacccg gggatcctct agagtcgacc tgcaggcatg
     1321 caagcttgcg gccgcgggcc cggtacctcg cgaaggcctt gcaggccaac cagataagtg
     1381 aaatctagtt ccaaactatt ttgtcatttt taattttcgt attagcttac gacgctacac
     1441 ccagttccca tctattttgt cactcttccc taaataatcc ttaaaaactc catttccacc
     1501 cctcccagtt cccaactatt ttgtccgccc acagcggggc atttttcttc ctgttatgtt
     1561 tgggcgctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     1621 tcttccgctt cctcgctcac tgacccgctg cgctcggtcg ttcggctgcg gcgagcggta
     1681 tcagagctta tcggccagcc tcgcagagca ggattcccgt tgagcaccgc caggtgcgaa
     1741 taagggacag tgaagaagga acacccgctc gcgggtgggc ctacttcacc tatcctgccc
     1801 ggctgacgcc gttggataca ccaaggaaag tctacacgaa ccctttggca aaatcctgta
     1861 tatcgtgcga aaaaggatgg atataccgaa aaaatcgcta taatgacccc gaagcagggt
     1921 tatgcagcgg aaagtatacc ttaaggaatc ccctgataac gcaggaaaga acatgtgagc
     1981 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgca ctccctgcct ctgtcatcac
     2041 gatactgtga tgccatggct aattcccatg tcagccgtta agtgttcctg tgtcactcaa
     2101 aattgctttg agaggctcta agggcttctc agtgcgttac atccctggct tgttgtccac
     2161 aaccgttaaa ccttaaaagc tttaaaagcc ttatatattc ttttttttct tataaaactt
     2221 aaaaccttag aggctattta agttgctgat ttatattaat tttattgttc aaacatgaga
     2281 gcttagtacg tgaaacatga gagcttagta cgttagccat gagagcttag tacgttagcc
     2341 atgagggttt agttcgttaa acatgagagc ttagtacgtt aaacatgaga gcttagtacg
     2401 tgaaacatga gagcttagta cgtactatca acaggttgaa ctgctgatct tcagatcctc
     2461 tacgccggac gcatcgtggc cggatcttgc ggccgcaaaa attaaaaatg aagttttaaa
     2521 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
     2581 gcaccaataa ctgcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     2641 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     2701 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     2761 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     2821 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     2881 tgctgcaatg ataccgcggg acccacgctc accggctcca gatttatcag caataaacca
     2941 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     3001 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     3061 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     3121 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     3181 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     3241 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     3301 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     3361 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     3421 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     3481 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     3541 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     3601 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     3661 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     3721 gcgcacattt ccccgaaaag tgccacctga cgtctaagaa accattatta tcatgacatt
     3781 aacctataaa aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg gtgacgacgg
     3841 tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc
     3901 cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct
     3961 taactatgcg gcatcagagc agattgtact gagagtgcac catatgcggt gtgaaatacc
     4021 gcacagatgc gtaaggagaa aataccgcat caggcgccat tcgccattca ggctgcgcaa
     4081 ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta cgccagctag aggaccagcc
     4141 gcgtaacctg gcaaaatcgg ttacggttga gtaataaatg gatgccctgc gtaagcaggt
     4201 gtgggcggac aataaagtct taaactgaac aaaatagatc taaactatga caataaagtc
     4261 ttaaactaga cagaatagtt gtaaactgaa atcagtccag ttatgctgtg aaaaagcata
     4321 ctggactttt gttatggcta aagcaaactc ttcattttct gaagtgcaaa ttgcccgtcg
     4381 tattaaagag gggcgtgggg ttcgaggtcg acggtatcga taagctagct taattagctg
     4441 agcttggact cctgttgata gatccagtaa tgacctcaga actccatctg gatttgttca
     4501 gaacgctcgg ttgccgccgg gcgtttttta ttggtgagaa tccaagctag actgcgatga
     4561 gtggcagggc ggggcgtaat ttttttaagg cagttattgg tgcccttaaa cgcctggggt
     4621 aatgactctc tagcttgagg catcaaataa aacgaaaggc tcagtcgaaa gactgggcct
     4681 ttcgttttat ctgttgtttg tcggtgaacg ctctcctgag taggacaaat ccgccgctag
     4741 gagcttgcgg cccggacgat gagctcgaat tggggatctt gaagtaccta ttccgaagtt
     4801 cctattctct agaaagtata ggaacttcag agcgcttttg aagctgatgc tcgagtgctt
     4861 aaaaacttac tcaatggaat aattctagat aattcttagg ccacacgttc aagtgcagcc
     4921 acaggataaa tttgcactga gcctgggtgg gattcggact cgaccgcata gccttcagga
     4981 gtgagttttg tgcaatacca accgacgact tgaccctgcc aagcggcacc agatttcttg
     5041 cgtacgcgat cccctaagcc aaaggtggca ctcaggggaa gcgcaaactg ccctgcaacg
     5101 ggagcgttgg cttcatcgct actttgaccc atgtcgaatc cttcttgtga atctattatg
     5161 gcgacaagca aatcgagctc tgactgccta ccccacaaca actatcagaa agcaccagca
     5221 caacggctgc ctaactttgt tttagggcga ctgccctgct gcgtaacatc gttgctgctc
     5281 cataacatca aacatcgacc cacggcgtaa cgcgcttgct gcttggatgc ccgaggcata
     5341 gactgtacaa aaaaacagtc ataacaagcc atgaaaaccg ccactgcgcc gttaccaccg
     5401 ctgcgttcgg tcaaggttct ggaccagttg cgtgagcgca tacgctactt gcattacagt
     5461 ttacgaaccg aacaggctta tgtcaactgg gttcgtgaat tatccattgc tgttgacaaa
     5521 gggaatcagg ggatcttgaa gttcctattc cgaagttcct attctctaga aagtatagga
     5581 acttcagagc gcttttgaag ctaattcgag ctcgatcatg catgagctca ctagtggcgc
     5641 g
//
