LOCUS       sequence-121748-        3852 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  source of CmR FRT cassette.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3852)
  AUTHORS   Choi KH, Gaynor JB, White KG, Lopez C, Bosio CM, Karkhoff-Schweizer 
            RR, Schweizer HP
  TITLE     A Tn7-based broad-range bacterial cloning and expression system.
  JOURNAL   Nat Methods. 2005 Jun;2(6):443-8.
  PUBMED    15908923
REFERENCE   2  (bases 1 to 3852)
  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 
            Methods."; date: "2005-06"; volume: "2(6)"; pages: "443-8"
COMMENT     Sequence Label: pFCM1
FEATURES             Location/Qualifiers
     source          1..3852
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     378..395
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    464..511
                     /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        596..698
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             699..1358
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(765..784)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     protein_bind    1498..1545
                     /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)."
     primer_bind     complement(1631..1647)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1631..1647)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1644..1666)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1655..1671
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(1679..1685,1686..1703,1704..1709))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1724..1745
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1862..1879)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2033..2621)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2113..2132)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2792..3583,3584..3652))
                     /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     3415..3434
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3653..3757)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3825..3843
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgccaa gcttgcatgc ctgcaggtcg
      421 actctagagg atccccgggt accgagctcg aattggggat cttgaagtac ctattccgaa
      481 gttcctattc tctagaaagt ataggaactt cagagcgctt ttgaagctga tgtgcttaaa
      541 aacttactca atggaataat tctagatggg ccaacttttg gcgaaaatga gacgttgatc
      601 ggcacgtaag aggttccaac tttcaccata atgaaataag atcactaccg ggcgtatttt
      661 ttgagttatc gagattttca ggagctaagg aagctaaaat ggagaaaaaa atcactggat
      721 ataccaccgt tgatatatcc caatggcatc gtaaagaaca ttttgaggca tttcagtcag
      781 ttgctcaatg tacctataac cagaccgttc agctggatat tacggccttt ttaaagaccg
      841 taaagaaaaa taagcacaag ttttatccgg cctttattca cattcttgcc cgcctgatga
      901 atgctcatcc ggagttccgt atggcaatga aagacggtga gctggtgata tgggatagtg
      961 ttcacccttg ttacaccgtt ttccatgagc aaactgaaac gttttcatcg ctctggagtg
     1021 aataccacga cgatttccgg cagtttctac acatatattc gcaagatgtg gcgtgttacg
     1081 gtgaaaacct ggcctatttc cctaaagggt ttattgagaa tatgtttttc gtctcagcca
     1141 atccctgggt gagtttcacc agttttgatt taaacgtggc caatatggac aacttcttcg
     1201 cccccgtttt caccatgggc aaatattata cgcaaggcga caaggtgctg atgccgctgg
     1261 cgattcaggt tcatcatgcc gtttgtgatg gcttccatgt cggcagaatg cttaatgaat
     1321 tacaacagta ctgcgatgag tggcagggcg gggcgtaatt tttttaaggc agttattggt
     1381 gcccttaaac gcctggttgc tacgcctgaa taagtgataa taagcggatg aatggcagaa
     1441 attcgaaagc aaattcgacc catccattgc tgttgacaaa gggaatcagg ggatcttgaa
     1501 gttcctattc cgaagttcct attctctaga aagtatagga acttcagagc gcttttgaag
     1561 ctaattcgag ctcggtaccc ggggatcctc tagagtcgac ctgcaggcat gcaagcttgg
     1621 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca
     1681 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca
     1741 cattaattgc gttgccctca ctgcccgctt tccagtcggg aagcctgtcg tgccagctgc
     1801 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     1861 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     1921 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     1981 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     2041 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     2101 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     2161 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     2221 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     2281 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     2341 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     2401 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     2461 gctacactag aaggacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     2521 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     2581 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     2641 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     2701 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     2761 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     2821 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     2881 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     2941 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     3001 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     3061 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     3121 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     3181 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     3241 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     3301 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     3361 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     3421 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     3481 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     3541 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     3601 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     3661 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
     3721 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     3781 ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga
     3841 ggccctttcg tc
//
