LOCUS       sequence-427294-        5283 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  High copy glmS-nadE plasmid containing chloramphenicol as antibiotic
            resistance gene (ARG) for validation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5283)
  AUTHORS   Amrofell MB, Rengarajan S, Vo ST, Ramirez Tovar ES, LoBello L, 
            Dantas G, Moon TS
  TITLE     Engineering E. coli strains using antibiotic-resistance-gene-free 
            plasmids.
  JOURNAL   Cell Rep Methods. 2023 Dec 18;3(12):100669. doi: 
            10.1016/j.crmeth.2023.100669. Epub 2023 Dec 11.
  PUBMED    38086386
REFERENCE   2  (bases 1 to 5283)
  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: "Cell Rep 
            Methods."; date: "2023-12-18"; volume: "3(12):100669. doi"; pages: "
            10.1016/j.crmeth.2023.100669. Epub 2023 Dec 11"
COMMENT     Sequence Label: pMBA326
FEATURES             Location/Qualifiers
     source          1..5283
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..35
                     /label=BBa_J23108
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.51"
     primer_bind     complement(2879..2896)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3050..3638)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3130..3149)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3809..4468)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     4383..4402
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(4469..4571)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     4672..4690
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ctgacagcta gctcagtcct aggtataatg ctagcgtcaa tttagtagta gaccacaagg
       61 aggttgctta atgtgtggaa ttgttggcgc gatcgcgcaa cgtgatgtag cagaaatcct
      121 tcttgaaggt ttacgtcgtc tggaataccg cggatatgac tctgccggtc tggccgttgt
      181 tgatgcagaa ggtcatatga cccgcctgcg tcgcctcggt aaagtccaga tgctggcaca
      241 ggcagcggaa gaacatcctc tgcatggcgg cactggtatt gctcacactc gctgggcgac
      301 ccacggtgaa ccttcagaag tgaatgcgca tccgcatgtt tctgaacaca ttgtggtggt
      361 gcataacggc atcatcgaaa accatgaacc gctgcgtgaa gagctaaaag cgcgtggcta
      421 taccttcgtt tctgaaaccg acaccgaagt gattgcccat ctggtgaact gggagctgaa
      481 acaaggcggg actctgcgtg aggccgttct gcgtgctatc ccgcagctgc gtggtgcgta
      541 cggtacagtg atcatggact cccgtcaccc ggataccctg ctggcggcac gttctggtag
      601 tccgctggtg attggcctgg ggatgggcga aaactttatc gcttctgacc agctggcgct
      661 gttgccggtg acccgtcgct ttatcttcct tgaagagggc gatattgcgg aaatcactcg
      721 ccgttcggta aacatcttcg ataaaactgg cgcggaagta aaacgtcagg atatcgaatc
      781 caatctgcaa tatgacgcgg gcgataaagg catttaccgt cactacatgc agaaagagat
      841 ctacgaacag ccgaacgcga tcaaaaacac ccttaccgga cgcatcagcc acggtcaggt
      901 tgatttaagc gagctgggac cgaacgccga cgaactgctg tcgaaggttg agcatattca
      961 gatcctcgcc tgtggtactt cttataactc cggtatggtt tcccgctact ggtttgaatc
     1021 gctagcaggt attccgtgcg acgtcgaaat cgcctctgaa ttccgctatc gcaaatctgc
     1081 cgtgcgtcgt aacagcctga tgatcacctt gtcacagtct ggcgaaaccg cggataccct
     1141 ggctggcctg cgtctgtcga aagagctggg ttaccttggt tcactggcaa tctgtaacgt
     1201 tccgggttct tctctggtgc gcgaatccga tctggcgcta atgaccaacg cgggtacaga
     1261 aatcggcgtg gcatccacta aagcattcac cactcagtta actgtgctgt tgatgctggt
     1321 ggcgaagctg tctcgcctga aaggtctgga tgcctccatt gaacatgaca tcgtgcatgg
     1381 tctgcaggcg ctgccgagcc gtattgagca gatgctgtct caggacaaac gcattgaagc
     1441 gctggcagaa gatttctctg acaaacatca cgcgctgttc ctgggccgtg gcgatcagta
     1501 cccaatcgcg ctggaaggcg cattgaagtt gaaagagatc tcttacattc acgctgaagc
     1561 ctacgctgct ggcgaactga aacacggtcc gctggcgcta attgatgccg atatgccggt
     1621 tattgttgtt gcaccgaaca acgaattgct ggaaaaactg aaatccaaca ttgaagaagt
     1681 tcgcgcgcgt ggcggtcagt tgtatgtctt cgccgatcag gatgcgggtt ttgtaagtag
     1741 cgataacatg cacatcatcg agatgccgca tgtggaagag gtgattgcac cgatcttcta
     1801 caccgttccg ctgcagctgc tggcttacca tgtcgcgctg atcaaaggca ccgacgttga
     1861 ccagccgcgt aacctggcaa aatcggttac ggttgagtaa gaacgatacc taatagaaaa
     1921 taggaattga caccaatgac attgcaacaa caaataataa aggcgctggg cgcaaaaccg
     1981 cagattaatg ctgaagagga aattcgtcgt agtatcgatt ttctgaaaag ctacctgcaa
     2041 acttatccgt tcattaaatc actggtgctc gggatcagcg gcggtcagga ctccacgctt
     2101 gccggtaagc tgtgccagat ggcgattaat gagctgcgcc aggaaaccgg caacgaatca
     2161 ctgcaattta ttgccgtacg cctgccctat ggtgtccagg ccgacgaaca agattgccag
     2221 gatgccatcg cctttattca accggatcgc gtattaaccg ttaatatcaa gggcgcggta
     2281 ctcgccagcg agcaggcatt gcgggaagca ggcattgaac tgagcgattt tgtccgtggc
     2341 aatgaaaaag cgcgtgagcg gatgaaagca caatatagca ttgcgggtat gaccagcggt
     2401 gtcgtggtgg gcaccgatca tgctgcagaa gctattaccg gattcttcac taaatatggt
     2461 gacggcggta cggacattaa tccgctgtat cgtctcaaca aacgtcaggg taaacagtta
     2521 ctggcggcat taggttgccc ggaacacctt tataagaaag cgccaacggc cgatcttgaa
     2581 gatgatcgcc cttctctgcc ggatgaagtg gcactcggcg tgacctatga caatatcgac
     2641 gactatctgg aagggaaaaa cgtacctgaa caggtcgcca gaacaataga gaactggtat
     2701 ctgaaaaccg aacataaacg ccgtccgcca attaccgttt tcgatgattt ctggaaaaag
     2761 taaagccgag caagacaatg gtaagtagta atagctcctg cccccggaag atcattccgg
     2821 gggctttttt attcccactg tgaactggat ccttactcga gtctagactg caggcttcct
     2881 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     2941 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     3001 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccacaggc
     3061 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     3121 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     3181 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     3241 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     3301 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     3361 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     3421 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     3481 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     3541 gagttggtag ctcttgatcc gccaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     3601 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     3661 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     3721 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     3781 gtatatatga gtaaacttgg tctgacagtt acgccccgcc ctgccactca tcgcagtact
     3841 gttgtaattc attaagcatt ctgccgacat ggaagccatc acaaacggca tgatgaacct
     3901 gaatcgccag cggcatcagc accttgtcgc cttgcgtata atatttgccc atggtgaaaa
     3961 cgggggcgaa gaagttgtcc atattggcca cgtttaaatc aaaactggtg aaactcaccc
     4021 agggattggc tgagacgaaa aacatattct caataaaccc tttagggaaa taggccaggt
     4081 tttcaccgta acacgccaca tcttgcgaat atatgtgtag aaactgccgg aaatcgtcgt
     4141 ggtattcact ccagagcgat gaaaacgttt cagtttgctc atggaaaacg gtgtaacaag
     4201 ggtgaacact atcccatatc accagctcac cgtctttcat tgccatacga aattccggat
     4261 gagcattcat caggcgggca agaatgtgaa taaaggccgg ataaaacttg tgcttatttt
     4321 tctttacggt ctttaaaaag gccgtaatat ccagctgaac ggtctggtta taggtacatt
     4381 gagcaactga ctgaaatgcc tcaaaatgtt ctttacgatg ccattgggat atatcaacgg
     4441 tggtatatcc agtgattttt ttctccattt tagcttcctt agctcctgaa aatctcgata
     4501 actcaaaaaa tacgcccggt agtgatctta tttcattatg gtgaaagttg gaacctctta
     4561 cgtgccgatc aacgtctcat tttcgccaga tatcgggttc cgcgcacatt tccccgaaaa
     4621 gtgccacctg acgtctaaga aaccattatt atcatgacat taacctataa aaataggcgt
     4681 atcacgaggc atcaccctag gtgtaaaacg acggccagac tcggtaccaa attccagaaa
     4741 agagacgctg aaaagcgtct tttttcgttt tggtccttat taatttttaa agtatgggct
     4801 taaccagatt agatcccgta gatgtccatc cccgccgaaa cgtttggtgg cgggaccagt
     4861 gacgaaggct tgagcgaggg cgtgcaagat tccgaatacc gcaagcgaca ggccgatcat
     4921 cgtcgcgctc cagcgaaagc ggtcctctcc gaaaatgacc cagagcgctg ccggcacctg
     4981 tcctacgagt tgcatgataa agaagacagt cataagtgcg gcgacgatag tcatgccccg
     5041 cgcccaccgg aaggagctga ctgggttgaa ggctctcaag ggcatcggtc gagatccggg
     5101 cttatcgact gcacggtgca ccaatgcttc tggcgtcagg cagccatcgg aagctgtggt
     5161 atggctgtgc aggtcgtaaa tcactgcata attcgtgtcg ctcaaggcgc actcccgttc
     5221 tggataatgt tttttgcgcc gacatcataa cggttctggc aaatattctg aaatgagctg
     5281 acc
//
