LOCUS       sequence-244278-        3522 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Variant of pKD13. Lambda Red recombineering PCR template plasmid 
            carrying an F5-tetA-F5 cassette orthogonal to FRT-KanR-FRT in pKD13.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3522)
  TITLE     Lambda Red recombineering PCR template plasmids, variants of pKD13 
            with orthogonal marker-FRT sites
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 3522)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pES133
FEATURES             Location/Qualifiers
     source          1..3522
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(125..1315)
                     /codon_start=1
                     /gene="tet"
                     /product="tetracycline efflux protein"
                     /label=TcR
                     /note="confers resistance to tetracycline"
                     /translation="MKSNNALIVILGTVTLDAVGIGLVMPVLPGLLRDIVHSDSIASHY
                     GVLLALYALMQFLCAPVLGALSDRFGRRPVLLASLLGATIDYAIMATTPVLWILYAGRI
                     VAGITGATGAVAGAYIADITDGEDRARHFGLMSACFGVGMVAGPVAGGLLGAISLHAPF
                     LAAAVLNGLNLLLGCFLMQESHKGERRPMPLRAFNPVSSFRWARGMTIVAALMTVFFIM
                     QLVGQVPAALWVIFGEDRFRWSATMIGLSLAVFGILHALAQAFVTGPATKRFGEKQAII
                     AGMAADALGYVLLAFATRGWMAFPIMILLASGGIGMPALQAMLSRQVDDDHQGQLQGSL
                     AALTSLTSITGPLIVTAIYAASASTWNGLAWIVGAALYLVCLPALRRGAWSRATST"
     primer_bind     944..963
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        complement(join(1363..1368,1369..1385,1386..1391))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     terminator      1516..1762
                     /label=lambda tL3 terminator
                     /note="transcription terminator tL3 from phage lambda"
     rep_origin      1823..2179
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     CDS             complement(join(2276..3067,3068..3136))
                     /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     2899..2918
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3137..3241)
                     /gene="bla"
                     /label=AmpR promoter
     terminator      3332..3418
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      join(3510..3522,1..15)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
ORIGIN
        1 gacggatggc ctttttgcgt ggccagtgcc aagcttgcat gcagattgca gcattacacg
       61 tcttgagcga ttgtgtaggc tggagctgct tcgaagttcc tatacctttt gaagaatagg
      121 aacttcaggt cgaggtggcc cggctccatg caccgcgacg caacgcgggg aggcagacaa
      181 ggtatagggc ggcgcctaca atccatgcca acccgttcca tgtgctcgcc gaggcggcat
      241 aaatcgccgt gacgatcagc ggtccagtga tcgaagttag gctggtaaga gccgcgagcg
      301 atccttgaag ctgtccctga tggtcgtcat ctacctgcct ggacagcatg gcctgcaacg
      361 cgggcatccc gatgccgccg gaagcgagaa gaatcataat ggggaaggcc atccagcctc
      421 gcgtcgcgaa cgccagcaag acgtagccca gcgcgtcggc cgccatgccg gcgataatgg
      481 cctgcttctc gccgaaacgt ttggtggcgg gaccagtgac gaaggcttga gcgagggcgt
      541 gcaagattcc gaataccgca agcgacaggc cgatcatcgt cgcgctccag cgaaagcggt
      601 cctcgccgaa aatgacccag agcgctgccg gcacctgtcc tacgagttgc atgataaaga
      661 atacagtcat aagtgcggcg acgatagtca tgccccgcgc ccaccggaag gagctgactg
      721 ggttgaaggc tctcaagggc atcggtcgac gctctccctt atgcgactcc tgcattagaa
      781 agcagcccag tagtaggttg aggccgttga gcaccgccgc cgcaaggaat ggtgcatgca
      841 aggagatggc gcccaacagt cccccggcca cggggcctgc caccataccc acgccgaaac
      901 aagcgctcat gagcccgaag tggcgagccc gatcttcccc atcggtgatg tcggcgatat
      961 aggcgccagc aaccgcacct gtggcgccgg tgatgccggc cacgatgcgt ccggcgtaga
     1021 gaatccacag gacgggtgtg gtcgccatga tcgcgtagtc gatagtggct ccaagtagcg
     1081 aagcgagcag gactgggcgg cggccaaagc ggtcggacag tgctccgaga acgggtgcgc
     1141 atagaaattg catcaacgca tatagcgcta gcagcacgcc atagtgactg gcgatgctgt
     1201 cggaatggac gatgtcccgc aagaggcccg gcagtaccgg cataaccaag cctatgccta
     1261 cagcatccag ggtgacggtg ccgaggatga cgatgagcgc attgttagat ttcatacacg
     1321 gtgcctgact gcgttagcaa tttaactgtg ataaactacc gcattaaagc ttatcgatga
     1381 taagctgtca aacatgagaa ttgaagttcc tatacctttt gaagaatagg aacttcgaac
     1441 tgcaggtcga cggatccccg gaattaattc tcatgtttga cagcttatca ctgatcagtg
     1501 aattaatggc gatgacgcat cctcacgata atatccgggt aggcgcaatc actttcgtct
     1561 ctactccgtt acaaagcgag gctgggtatt tcccggcctt tctgttatcc gaaatccact
     1621 gaaagcacag cggctggctg aggagataaa taataaacga ggggctgtat gcacaaagca
     1681 tcttctgttg agttaagaac gagtatcgag atggcacata gccttgctca aattggaatc
     1741 aggtttgtgc caataccagt agaaacagac gaagaagcta gctttgcact ggattgcgag
     1801 gctttgccat ggctaattcc catgtcagcc gttaagtgtt cctgtgtcac tgaaaattgc
     1861 tttgagaggc tctaagggct tctcagtgcg ttacatccct ggcttgttgt ccacaaccgt
     1921 taaaccttaa aagctttaaa agccttatat attctttttt ttcttataaa acttaaaacc
     1981 ttagaggcta tttaagttgc tgatttatat taattttatt gttcaaacat gagagcttag
     2041 tacgtgaaac atgagagctt agtacgttag ccatgagagc ttagtacgtt agccatgagg
     2101 gtttagttcg ttaaacatga gagcttagta cgttaaacat gagagcttag tacgtgaaac
     2161 atgagagctt agtacgtact atcaacaggt tgaactgcgg atcttgcggc cgcaaaaatt
     2221 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     2281 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     2341 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     2401 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     2461 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     2521 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     2581 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     2641 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     2701 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     2761 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     2821 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     2881 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     2941 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     3001 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     3061 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     3121 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     3181 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     3241 gcacatttcc ccgaaaagtg ccacctgcat cgatggcccc ccgatggtag tgtggggtct
     3301 ccccatgcga gagtagggaa ctgccaggca tcaaataaaa cgaaaggctc agtcgaaaga
     3361 ctgggccttt cgttttatct gttgtttgtc ggtgaacgct ctcctgagta ggacaaatcc
     3421 gccgggagcg gatttgaacg ttgcgaagca acggcccgga gggtggcggg caggacgccc
     3481 gccataaact gccaggcatc aaattaagca gaaggccatc ct
//
