LOCUS       sequence-468004-        6281 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CCP expression in diverse gram-negative bacteria, with pre-cloned 
            Golden Gate Compatible (BsaI) restriction sites to facilitate 
            promoter swapping. .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 3688 to 3722)
  AUTHORS   Armstrong ZR, Alonso J, Stanton V, Patel N, Zogaj X, Cocioba SS, 
            Klose KE
  TITLE     Mobilizable shuttle vectors with fluorescent markers functional 
            across different species of bacteria.
  JOURNAL   Appl Environ Microbiol. 2025 May 12:e0004525. doi: 
            10.1128/aem.00045-25.
  PUBMED    40353662
REFERENCE   2  (bases 1 to 6281)
  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: "Appl 
            Environ Microbiol. 2025 May 12:e0004525. doi: 10.1128/aem.00045-25."
COMMENT     Sequence Label: pKEK3221
FEATURES             Location/Qualifiers
     source          1..6281
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     oriT            924..1033
                     /note="incP origin of transfer"
     CDS             1066..1437
                     /codon_start=1
                     /gene="traJ"
                     /product="oriT-recognizing protein"
                     /label=traJ
                     /translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSLSAYLLAV
                     GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
                     KQDELGKVMMGVVRPRAEP"
     CDS             complement(2042..2719)
                     /codon_start=1
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGRKRR
                     ADPA"
     primer_bind     2634..2653
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     primer_bind     2911..2927
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2911..2927
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     RBS             3664..3675
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     promoter        complement(3688..3722)
                     /label=﻿BBa_J23100
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 1.0
                     "
                     /citation=[1]
     primer_bind     complement(3729..3745)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      3799..4344
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     4245..4264
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4461..4478
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 aaaaaaaaat ctgacaatat ataaagagaa tatataaaga gaatatctta ggggatttta
       61 aaaaaatccc acagactcaa agactttttt gactttttaa atcctagaaa ctatacttta
      121 agtacttatt taagtacatg gatttagatt atgcaaaccg ttaattattc aacttttaga
      181 aatgaactat ctgattcaat ggatagagta acaaaaaatc atagtcctat gattgtaact
      241 agaggttcaa aaaaagaagc agttgttatg atgtcgttag aggattttaa agctttgtta
      301 gcagcctttt tgtccggtgt tgggttgaag gtgaagccgg tcggggccgc agcgggggcc
      361 ggcttttcag ccttgccccc ctgcttcggc cgccgtggct ccggcgtctt gggtgccggc
      421 gcgggttccg cagccttggc ctgcggtgcg ggcacatcgg cgggcttggc cttgatgtgc
      481 cgcctggcgt gcgagcggaa cgtctcgtag gagaacttga ccttccccgt ttcccgcatg
      541 tgctcccaaa tggtgacgag cgcatagccg gacgctaacg ccgcctcgac atccgccctc
      601 accgccagga acgcaaccgc agcctcatca cgccggcgct tcttggccgc gcgggattca
      661 acccactcgg ccagctcgtc ggtgtagctc tttggcatcg tctctcgcct gtcccctcag
      721 ttcagtaatt tcctgcattt gcctgtttcc agtcggtaga tattccacaa aacagcaggg
      781 aagcagcgct tttccgctgc ataaccctgc ttcggggtca ttatagcgat tttttcggta
      841 tatccatcct ttttcgcacg atatacagga ttttgccaaa gggttcgtgt agactttcct
      901 tggtgtatcc aacggcgtca gccgggcagg ataggtgaag taggcccacc cgcgagcggg
      961 tgttccttct tcactgtccc ttattcgcac ctggcggtgc tcaacgggaa tcctgctctg
     1021 cgaggctggc cggctaccgc cggcgtaaca gatgagggca agcggatggc tgatgaaacc
     1081 aagccaacca ggaagggcag cccacctatc aaggtgtact gccttccaga cgaacgaaga
     1141 gcgattgagg aaaaggcggc ggcggccggc atgagcctgt cggcctacct gctggccgtc
     1201 ggccagggct acaaaatcac gggcgtcgtg gactatgagc acgtccgcga gctggcccgc
     1261 atcaatggcg acctgggccg cctgggcggc ctgctgaaac tctggctcac cgacgacccg
     1321 cgcacggcgc ggttcggtga tgccacgatc ctcgccctgc tggcgaagat cgaagagaag
     1381 caggacgagc ttggcaaggt catgatgggc gtggtccgcc cgagggcaga gccatgactt
     1441 ttttagccgc taaaacggcc ggggggtgcg cgtgattgcc aagcacgtcc ccatgcgctc
     1501 catcaagaag agcgacttcg cggagctggt gaagtacatc accgacgagc aaggcaagac
     1561 cgagcgcctg ggtcacgtgc gcgtcacgaa ctgcgaggca aacaccctgc ccgctgtcat
     1621 ggccgaggtg atggcgaccc agcacggcaa cacccgttcc gaggccgaca agacctatca
     1681 cctcctggtt agcttccgcg cgggagagaa gcccgacgcg gagacgttgc gcgcgattga
     1741 ggaccgcatc tgcgctgggc ttggcttcgc cgagcatcag cgcgtcagtg ccgtgcatca
     1801 cgacaccgac aacctgcaca tccatatcgc catcaacaag attcacccga cccgaaacac
     1861 catccatgag ccgtatcggg cctaccgcgc cctcgctgac ctctgcgcga cgctcgaacg
     1921 ggactacggg cttgagcgtg acaatcacga aacgcggcag cgcgtttccg agaaccgcgc
     1981 gaacgacatg gagcggcacg cgggcgtgga aagcctggtc ggctgctcgc gctttgttag
     2041 cagccggatc cgcgcggcgt ttacgccccc cctgccactc atcgcagtac tgttgtaatt
     2101 cattaagcat tctgccgaca tggaagccat cacaaacggc atgatgaacc tgaatcgcca
     2161 gcggcatcag caccttgtcg ccttgcgtat aatatttgcc catggtgaaa acgggggcga
     2221 agaagttgtc catattggcc acgtttaaat caaaactggt gaaactcacc cagggattgg
     2281 ctgagacgaa aaacatattc tcaataaacc ctttagggaa ataggccagg ttttcaccgt
     2341 aacacgccac atcttgcgaa tatatgtgta gaaactgccg gaaatcgtcg tggtattcac
     2401 tccagagcga tgaaaacgtt tcagtttgct catggaaaac ggtgtaacaa gggtgaacac
     2461 tatcccatat caccagctca ccgtctttca ttgccatacg gaattccgga tgagcattca
     2521 tcaggcgggc aagaatgtga ataaaggccg gataaaactt gtgcttattt ttctttacgg
     2581 tctttaaaaa ggccgtaata tccagctgaa cggtctggtt ataggtacat tgagcaactg
     2641 actgaaatgc ctcaaaatgt tctttacgat gccattggga tatatcaacg gtggtatatc
     2701 cagtgatttt tttctccata tgtttagtct cctttttttg ttataaatat tttatacaca
     2761 aaaatcttcg ttatttaaat aagttttata gctatttttt caagatacaa aactattaat
     2821 tagttataat taggttatta gcttaaaatt ataaaccaaa tacgatgagt gacaacccaa
     2881 aagatctcga tcctctacgc cgttggggag caggaaacag ctatgacggt accaaaaaaa
     2941 accccgccct gtcaggggcg gggttttttt ttgagctctt atcatgtaga gtgacgtgcc
     3001 tccgcgcgct cgtattgctc aacaatggtg tagtcctcgt tatgtgacgt aatatctaac
     3061 ttaatgtcca cgttataggc cccgggcaat tgcacgggct tcttggcttt gtaagtagtt
     3121 ttgacttccg cgtcatagtg tccaccatcc ttgagcttga gtcgttggtt gatttcaccc
     3181 ttcaacgcac cgtcctctgg gtacatccgt tccgtactag cttcccatcc catagtcttc
     3241 ttctgcatca ctggtccatc agaagggaaa ttaatgccga gcagcttaac tttgtagatg
     3301 aactgaccat cttgcagaga agagtcctgg gttaccgtga ctacaccgcc gtcttcaaaa
     3361 ttcattacac gctcccaggt gaatccctct ggaaaggaaa gtttcaaata gtccggaata
     3421 tcagctggat gctttacata cgctttcgaa ccatacataa actgaggact taagatatcc
     3481 cacgcgaacg gcagtggacc gcccttagtt actttcagct tggcggtttg agttccttcg
     3541 taggggtggc cttcaccctc accttcgatc tcgaactcat gaccgttgac agacccttcc
     3601 atgtggacct taaaacgcat aaattccttg ataattgcca tattatcttc ttccatggga
     3661 ctctttctcc tctttaatcg agaccaagct agcactgtac ctaggactga gctagccgtc
     3721 aagaattcac tggccgtcgt tttacggtct cagtcgacta ctctagaaat attttatctg
     3781 attaataaga tgatcttctt gagatcgttt tggtctgcgc gtaatctctt gctctgaaaa
     3841 cgaaaaaacc gccttgcagg gcggtttttc gaaggttctc tgagctacca actctttgaa
     3901 ccgaggtaac tggcttggag gagcgcagtc accaaaactt gtcctttcag tttagcctta
     3961 accggcgcat gacttcaaga ctaactcctc taaatcaatt accagtggct gctgccagtg
     4021 gtgcttttgc atgtctttcc gggttggact caagacgata gttaccggat aaggcgcagc
     4081 ggtcggactg aacggggggt tcgtgcatac agtccagctt ggagcgaact gcctacccgg
     4141 aactgagtgt caggcgtgga atgagacaaa cgcggccata acagcggaat gacaccggta
     4201 aaccgaaagg caggaacagg agagcgcacg agggagccgc cagggggaaa cgcctggtat
     4261 ctttatagtc ctgtcgggtt tcgccaccac tgatttgagc gtcagatttc gtgatgcttg
     4321 tcaggggggc ggagcctatg gaaaaacggc tttgccgcgg ccctctcact tccctgttaa
     4381 gtatcttcct ggcatcttcc aggaaatctc cgccccgttc gtaagccatt tccgctcgcc
     4441 gcagtcgaac gaccgagcgt agcgagtcag tgagcgagga agcggaatat atcctgtatc
     4501 acatattctg ctgacgcacc ggtgcagcct tttttctcct gccacatgaa gcacttcact
     4561 gacaccctca tcagtgccaa catagtaagc cagtatacac tccgctagcg ctgatgtccg
     4621 gcggtgcttt tgccgttacg caccaccccg tcagtagctg aacaggaggg acagctaaag
     4681 ctagaaatgt accaaatttt acctatttta ggatactttt catgaagctc ttctattttt
     4741 ccccagtcct ttaatagtct acctttagag tctcgtaaat agttatcttt gtgacagggg
     4801 cctcttttat cttttttaat gtaactatat gttattccaa cgtcactatt actattatcc
     4861 aaatcttttt tagcatgcca gtaagaactt tcataactta actctatctt tcgacctctt
     4921 tgatatacaa caataaagct atagccagta gtaacaacct gttttacttt tgttaaatct
     4981 attaacttct tatttatttt tttatgtttt tttgaaaatt taaatatttc tatattcatt
     5041 cctacacttc ctcaaatcca aatggtagct tatgattctc ttctggtttc ttttctaatt
     5101 tttttatatt tgcaataaaa actctttttc tatctttgat ttttttattg tcccaattcc
     5161 tccaagtatc atcacaaacc ctttcaatat catgtaaatg atgatgtcta aatattgatc
     5221 tgacataata cagatctagg tctagttcat cacttaacac aacttctcta agtctttcag
     5281 atgcttcgat tggtatgtaa tcctctttat ttttagtatc taaaggcttt tgcttaaatt
     5341 cttcttctgt ctctgctacc ttactaactg taaacttgat atttgtaatc ttacgaccat
     5401 gttttctgtg atgatccttg tcatcatagg ttacaaaaat atccgataat tgattaatct
     5461 cttctagtgc tggtaatagg aacttatttt taaaatttga atatctgttg ctgtaacttt
     5521 taggtaaatc aaaatcatta atcatatcat cgacatacaa tacgcaatca actatattag
     5581 cataccctgc ttgttcgcct aatttgcttt tgagaagtaa gtataatctg cttgaatact
     5641 tacttttaaa tgaaaatagt aactgtcttt ctgctttagt aaagtactct tgtagttgta
     5701 tcatgtgtgg cattaatgac caatgaaact cgcaaattaa agcactgctt ttagggtctg
     5761 cttcaatata tgcaaaccag ttagctatct tcgtttgttc tttattcagc catactggct
     5821 tagacattat tgagtgcatt aattgcttca atctcactct gttatgctta acccctgtag
     5881 ctttttcaag atcagatagg cttatcttat acctgtgaaa ctctttatct tctcttttaa
     5941 ccattgaggc aactaagaat attaagtttt gttcttcttt tgtaaggcta tactttcctg
     6001 caacaagagt attagacata gctatttctt tgccagcatt tacattttta acttctttca
     6061 tagaactaga gtcattatct cgatatacaa attctataaa acttctatta gtaaaacaac
     6121 tacttcataa aaaaaagtag ttttaacgat acaaaaagta gttttaaatt caaaaagtga
     6181 tacaaaaagt agttttaaat tcaaaaagtg atacaaaaag tagttttaaa ttttttaaaa
     6241 aagtgcttca aagccttatg tagcaatact tacagaggat t
//
