LOCUS       sequence-260072-        7312 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Arabinose inducible expression of lamba-Red recombinase proteins and
            I-CreI..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7312)
  AUTHORS   Wei Y, Deng P, Mohsin A, Yang Y, Zhou H, Guo M, Fang H
  TITLE     An electroporation-free method based on Red recombineering for 
            markerless deletion and genomic replacement in the Escherichia coli 
            DH1 genome.
  JOURNAL   PLoS One. 2017 Oct 24;12(10):e0186891. doi: 
            10.1371/journal.pone.0186891. eCollection 2017.
  PUBMED    29065183
REFERENCE   2  (bases 1 to 7312)
  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: "PLoS One.";
            date: "2017-10-24"; volume: "12(10):e0186891. doi"; pages: " 
            10.1371/journal.pone.0186891. eCollection 2017"
COMMENT     Sequence Label: pCNA
FEATURES             Location/Qualifiers
     source          1..7312
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             138..554
                     /codon_start=1
                     /gene="gam"
                     /product="inhibitor of the host RecBCD nuclease in the 
                     lambda Red system"
                     /label=Gam
                     /note="protects the double-stranded DNA substrate for 
                     recombination"
                     /translation="MDINTETEIKQKHSLTPFPVFLISPAFRGRYFHSYFRSSAMNAYY
                     IQDRLEAQSWARHYQQLAREEKEAELADDMEKGLPQHLFESLCIDHLQRHGASKKSITR
                     AFDDDVEFQERMAEHIRYMVETIAHHQVDIDSEV"
     CDS             560..1345
                     /codon_start=1
                     /gene="bet"
                     /product="single-stranded DNA binding recombinase in the 
                     lambda Red system"
                     /label=Beta
                     /note="mediates pairing of a single-stranded DNA with a 
                     complementary target"
                     /translation="MSTALATLAGKLAERVGMDSVDPQELITTLRQTAFKGDASDAQFI
                     ALLIVANQYGLNPWTKEIYAFPDKQNGIVPVVGVDGWSRIINENQQFDGMDFEQDNESC
                     TCRIYRKDRNHPICVTEWMDECRREPFKTREGREITGPWQSHPKRMLRHKAMIQCARLA
                     FGFAGIYDKDEAERIVENTAYTAERQPERDITPVNDETMQEINTLLIALDKTWDDDLLP
                     LCSQIFRRDIRASSELTQAEAVKALGFLKQKAAEQKVAA"
     CDS             1342..2022
                     /codon_start=1
                     /gene="exo"
                     /product="5' to 3' double-stranded DNA exonuclease in the 
                     lambda Red system"
                     /label=Exo
                     /note="creates 3' single-stranded DNA tails"
                     /translation="MTPDIILQRTGIDVRAVEQGEDAWHKLRLGVITASEVHNVIAKPR
                     SGKKWPDMKMSYFHTLLAEVCTGVAPEVNAKALAWGKQYENDARTLFEFTSGVNVTESP
                     IIYRDESMRTACSPDGLCSDGNGLELKCPFTSRDFMKFRLGGFEAIKSAYMAQVQYSMW
                     VTRKNAWYFANYDPRMKREGLHYVVIERDEKYMASFDEIVPEFIEKMDEALAEIGFVFG
                     EQWR"
     CDS             complement(2520..3470)
                     /codon_start=1
                     /gene="rep101 encoding an A56V mutation"
                     /product="temperature-sensitive version of the RepA protein
                     needed for replication with the pSC101 origin (Armstrong et
                     al., 1984)"
                     /label=Rep101(Ts)
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYVQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
     rep_origin      3518..3740
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     primer_bind     complement(4333..4351)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4419..4523
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4524..4592,4593..5384)
                     /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     complement(4742..4761)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     CDS             complement(5704..6582)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     promoter        6609..6893
                     /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     6816..6835
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
ORIGIN
        1 gattctaaga ctcgtaaaac cacttctgaa accgttcgtg ctgtgctgga cagcctgagc
       61 gagaagaaga aatcctcccc ggcggccgac taatctagcg aaaagatgtt tcgtgaagcc
      121 gtcgacgctt ataaaaaatg gatattaata ctgaaactga gatcaagcaa aagcattcac
      181 taaccccctt tcctgttttc ctaatcagcc cggcatttcg cgggcgatat tttcacagct
      241 atttcaggag ttcagccatg aacgcttatt acattcagga tcgtcttgag gctcagagct
      301 gggcgcgtca ctaccagcag ctcgcccgtg aagagaaaga ggcagaactg gcagacgaca
      361 tggaaaaagg cctgccccag cacctgtttg aatcgctatg catcgatcat ttgcaacgcc
      421 acggggccag caaaaaatcc attacccgtg cgtttgatga cgatgttgag tttcaggagc
      481 gcatggcaga acacatccgg tacatggttg aaaccattgc tcaccaccag gttgatattg
      541 attcagaggt ataaaacgaa tgagtactgc actcgcaacg ctggctggga agctggctga
      601 acgtgtcggc atggattctg tcgacccaca ggaactgatc accactcttc gccagacggc
      661 atttaaaggt gatgccagcg atgcgcagtt catcgcatta ctgatcgttg ccaaccagta
      721 cggccttaat ccgtggacga aagaaattta cgcctttcct gataagcaga atggcatcgt
      781 tccggtggtg ggcgttgatg gctggtcccg catcatcaat gaaaaccagc agtttgatgg
      841 catggacttt gagcaggaca atgaatcctg tacatgccgg atttaccgca aggaccgtaa
      901 tcatccgatc tgcgttaccg aatggatgga tgaatgccgc cgcgaaccat tcaaaactcg
      961 cgaaggcaga gaaatcacgg ggccgtggca gtcgcatccc aaacggatgt tacgtcataa
     1021 agccatgatt cagtgtgccc gtctggcctt cggatttgct ggtatctatg acaaggatga
     1081 agccgagcgc attgtcgaaa atactgcata cactgcagaa cgtcagccgg aacgcgacat
     1141 cactccggtt aacgatgaaa ccatgcagga gattaacact ctgctgatcg ccctggataa
     1201 aacatgggat gacgacttat tgccgctctg ttcccagata tttcgccgcg acattcgtgc
     1261 atcgtcagaa ctgacacagg ccgaagcagt aaaagctctt ggattcctga aacagaaagc
     1321 cgcagagcag aaggtggcag catgacaccg gacattatcc tgcagcgtac cgggatcgat
     1381 gtgagagctg tcgaacaggg ggaagatgcg tggcacaaat tacggctcgg cgtcatcacc
     1441 gcttcagaag ttcacaacgt gatagcaaaa ccccgctccg gaaagaagtg gcctgacatg
     1501 aaaatgtcct acttccacac cctgcttgct gaggtttgca ccggtgtggc tccggaagtt
     1561 aacgctaaag cactggcctg gggaaaacag tacgagaacg acgccagaac cctgtttgaa
     1621 ttcacttccg gcgtgaatgt tactgaatcc ccgatcatct atcgcgacga aagtatgcgt
     1681 accgcctgct ctcccgatgg tttatgcagt gacggcaacg gccttgaact gaaatgcccg
     1741 tttacctccc gggatttcat gaagttccgg ctcggtggtt tcgaggccat aaagtcagct
     1801 tacatggccc aggtgcagta cagcatgtgg gtgacgcgaa aaaatgcctg gtactttgcc
     1861 aactatgacc cgcgtatgaa gcgtgaaggc ctgcattatg tcgtgattga gcgggatgaa
     1921 aagtacatgg cgagttttga cgagatcgtg ccggagttca tcgaaaaaat ggacgaggca
     1981 ctggctgaaa ttggttttgt atttggggag caatggcgat gacgcatcct cacgataata
     2041 agcttcctgc tgaacatcaa aggcaagaaa acatctgttg tcaaagacag catccttgaa
     2101 caaggacaat taacagttaa caaataaaaa cgcaaaagaa aatgccgata ttgactaccg
     2161 gaagcagtgt gaccgtgtgc ttctcaaatg cctgattcag gctgtctatg tgtgactgtt
     2221 gagctgtaac aagttgtctc aggtgttcaa tttcatgttc tagttgcttt gttttactgg
     2281 tttcacctgt tctattaggt gttacatgct gttcatctgt tacattgtcg atctgttcat
     2341 ggtgaacagc tttaaatgca ccaaaaactc gtaaaagctc tgatgtatct atctttttta
     2401 caccgttttc atctgtgcat atggacagtt ttccctttga tatgtaacgg tgaacagttg
     2461 ttctactttt gtttgttagt cttgatgctt cactgataga tacaagagcc ataagaacct
     2521 cagatccttc cgtatttagc cagtatgttc tctagtgtgg ttcgttgttt ttgcgtgagc
     2581 catgagaacg aaccattgag atcatactta ctttgcatgt cactcaaaaa ttttgcctca
     2641 aaactggtga gctgaatttt tgcagttaaa gcatcgtgta gtgtttttct tagtccgtta
     2701 tgtaggtagg aatctgatgt aatggttgtt ggtattttgt caccattcat ttttatctgg
     2761 ttgttctcaa gttcggttac gagatccatt tgtctatcta gttcaacttg gaaaatcaac
     2821 gtatcagtcg ggcggcctcg cttatcaacc accaatttca tattgctgta agtgtttaaa
     2881 tctttactta ttggtttcaa aacccattgg ttaagccttt taaactcatg gtagttattt
     2941 tcaagcatta acatgaactt aaattcatca aggctaatct ctatatttgc cttgtgagtt
     3001 ttcttttgtg ttagttcttt taataaccac tcataaatcc tcatagagta tttgttttca
     3061 aaagacttaa catgttccag attatatttt atgaattttt ttaactggaa aagataaggc
     3121 aatatctctt cactaaaaac taattctaat ttttcgcttg agaacttggc atagtttgtc
     3181 cactggaaaa tctcaaagcc tttaaccaaa ggattcctga tttccacagt tctcgtcatc
     3241 agctctctgg ttgctttagc taatacacca taagcatttt ccctactgat gttcatcatc
     3301 tgaacgtatt ggttataagt gaacgatacc gtccgttctt tccttgtagg gttttcaatc
     3361 gtggggttga gtagtgccac acagcataaa attagcttgg tttcatgctc cgttaagtca
     3421 tagcgactaa tcgctagttc atttgctttg aaaacaacta attcagacat acatctcaat
     3481 tggtctaggt gattttaatc actataccaa ttgagatggg ctagtcaatg ataattacta
     3541 gtccttttcc tttgagttgt gggtatctgt aaattctgct agacctttgc tggaaaactt
     3601 gtaaattctg ctagaccctc tgtaaattcc gctagacctt tgtgtgtttt ttttgtttat
     3661 attcaagtgg ttataattta tagaataaag aaagaataaa aaaagataaa aagaatagat
     3721 cccagccctg tgtataactc actactttag tcagttccgc agtattacaa aaggatgtcg
     3781 caaacgctgt ttgctcctct acaaaacaga ccttaaaacc ctaaaggctt aagtagcacc
     3841 ctcgcaagct cgggcaaatc gctgaatatt ccttttgtct ccgaccatca ggcacctgag
     3901 tcgctgtctt tttcgtgaca ttcagttcgc tgcgctcacg gctctggcag tgaatggggg
     3961 taaatggcac tacaggcgcc ttttatggat tcatgcaagg aaactaccca taatacaaga
     4021 aaagcccgtc acgggcttct cagggcgttt tatggcgggt ctgctatgtg gtgctatctg
     4081 actttttgct gttcagcagt tcctgccctc tgattttcca gtctgaccac ttcggattat
     4141 cccgtgacag gtcattcaga ctggctaatg cacccagtaa ggcagcggta tcatcaacag
     4201 gcttacccgt cttactgtcg gggatcgacg ctctccctta tgcgactcct gcacctttcg
     4261 tcttcgaata aatacctgtg acggaagatc acttcgcaga ataaataaat cctggtgtcc
     4321 ctggacgaaa gggcctcgtg atacgcctat ttttataggt taatgtcatg ataataatgg
     4381 tttcttagac gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     4441 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     4501 aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct
     4561 tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag
     4621 atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta
     4681 agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc
     4741 tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca
     4801 tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg
     4861 atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg
     4921 ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca
     4981 tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa
     5041 acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa
     5101 ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg gaggcggata
     5161 aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat
     5221 ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc
     5281 cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata
     5341 gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt
     5401 actcatatat actttagatt gcgtaatttt tttaaggcag ttattggtgc ccttaaacgc
     5461 ctggttgcta cgcctgaata agtgataata agcggatgaa tggcagaaat tcgaaagcaa
     5521 attcgacccg gtcgtcggtt cagggcaggg tcgttaaata gccgcttatg tctattgctg
     5581 gtctcggtac ccggggatcc tctagagtcg acctgcataa tgtgcctgtc aaatggacga
     5641 agcagggatt ctgcaaaccc tatgctactc cgtcaagccg tcaattgtct gattcgttac
     5701 caattatgac aacttgacgg ctacatcatt cactttttct tcacaaccgg cacggaactc
     5761 gctcgggctg gccccggtgc attttttaaa tacccgcgag aaatagagtt gatcgtcaaa
     5821 accaacattg cgaccgacgg tggcgatagg catccgggtg gtgctcaaaa gcagcttcgc
     5881 ctggctgata cgttggtcct cgcgccagct taagacgcta atccctaact gctggcggaa
     5941 aagatgtgac agacgcgacg gcgacaagca aacatgctgt gcgacgctgg cgatatcaaa
     6001 attgctgtct gccaggtgat cgctgatgta ctgacaagcc tcgcgtaccc gattatccat
     6061 cggtggatgg agcgactcgt taatcgcttc catgcgccgc agtaacaatt gctcaagcag
     6121 atttatcgcc agcagctccg aatagcgccc ttccccttgc ccggcgttaa tgatttgccc
     6181 aaacaggtcg ctgaaatgcg gctggtgcgc ttcatccggg cgaaagaacc ccgtattggc
     6241 aaatattgac ggccagttaa gccattcatg ccagtaggcg cgcggacgaa agtaaaccca
     6301 ctggtgatac cattcgcgag cctccggatg acgaccgtag tgatgaatct ctcctggcgg
     6361 gaacagcaaa atatcacccg gtcggcaaac aaattctcgt ccctgatttt tcaccacccc
     6421 ctgaccgcga atggtgagat tgagaatata acctttcatt cccagcggtc ggtcgataaa
     6481 aaaatcgaga taaccgttgg cctcaatcgg cgttaaaccc gccaccagat gggcattaaa
     6541 cgagtatccc ggcagcaggg gatcattttg cgcttcagcc atacttttca tactcccgcc
     6601 attcagagaa gaaaccaatt gtccatattg catcagacat tgccgtcact gcgtctttta
     6661 ctggctcttc tcgctaacca aaccggtaac cccgcttatt aaaagcattc tgtaacaaag
     6721 cgggaccaaa gccatgacaa aaacgcgtaa caaaagtgtc tataatcacg gcagaaaagt
     6781 ccacattgat tatttgcacg gcgtcacact ttgctatgcc atagcatttt tatccataag
     6841 attagcggat cctacctgac gctttttatc gcaactctct actgtttctc catacccgtt
     6901 tttttgggct agctaagaag gagatataca tatgttatac ctggcaggtt tcgtggacgg
     6961 tgacggtagc atcatcgctc agattaaacc aaaccagtct tataagttta aacatcagct
     7021 gagcttgacc tttcaggtga ctcaaaagac ccagcgccgt tggtttctgg acaaactagt
     7081 ggatgaaatt ggcgttggtt acgtacgtga tcgcggttcc gtttccgatt acatcttaag
     7141 cgaaatcaag ccgctgcaca acttcctgac tcaactgcag ccgtttctga aactgaaaca
     7201 gaaacaggca aacctggttc tgaaaattat cgaacagctg ccgtctgcaa aagaatcccc
     7261 ggacaaattc ctggaagttt gtacctgggt ggatcagatt gcagctctga ac
//
