LOCUS       40924_27563        6812 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Gene replacement vector pLAW344, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6812)
  AUTHORS   Edelstein PH, Hu B.
  TITLE     Direct Submission
  JOURNAL   
REFERENCE   2  (bases 1 to 6812)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 6812)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Submitted 
            (14-NOV-2005) Pathology "
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..6812
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(208..1065)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(1066..1170)
                     /label=AmpR promoter
     protein_bind    1301..1334
                     /label=loxP
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(1394..2812)
                     /codon_start=1
                     /label=SacB
                     /note="secreted levansucrase that renders bacterial growth 
                     sensitive to sucrose"
                     /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
                     ITRHDMLQIPEQQKNEKYQVPEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYH
                     IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
                     SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
                     KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
                     YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
                     IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
                     MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
                     VKDSILEQGQLTVNK"
     promoter        complement(2813..3258)
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     primer_bind     complement(3333..3349)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     oriT            3722..3831
                     /label=oriT
                     /note="incP origin of transfer"
     CDS             3864..4232
                     /codon_start=1
                     /label=traJ
                     /note="oriT-recognizing protein"
                     /translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSLSAYLLAV
                     GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
                     KQDELGKVMMGVVRPRAEP"
     promoter        4705..4807
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding
                     chloramphenicol acetyltransferase"
     CDS             4808..5464
                     /codon_start=1
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     misc_feature    5932..6072
                     /label=bom
                     /note="basis of mobility region from pBR322"
     rep_origin      complement(join(6258..6812,1..34))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
       61 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
      121 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
      181 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
      241 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
      301 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
      361 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
      421 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
      481 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct gcaggcatcg tggtgtcacg
      541 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
      601 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
      661 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
      721 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
      781 atagtgtatg cggcgaccga gttgctcttg cccggcgtca acacgggata ataccgcgcc
      841 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
      901 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
      961 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     1021 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
     1081 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
     1141 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt
     1201 ctaagaaacc attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt
     1261 tcgtcttcaa gaattccgat catattcaat aacccttaat ataacttcgt ataatgtatg
     1321 ctatacgaag ttattaggtc tgaagaggag tttacgtcca gccaagctat cggcattttc
     1381 ttttgcgttt ttatttgtta actgttaatt gtccttgttc aaggatgctg tctttgacaa
     1441 cagatgtttt cttgcctttg atgttcagca ggaagcttgg cgcaaacgtt gattgtttgt
     1501 ctgcgtagaa tcctctgttt gtcatatagc ttgtaatcac gacattgttt cctttcgctt
     1561 gaggtacagc gaagtgtgag taagtaaagg ttacatcgtt aggatcaaga tccattttta
     1621 acacaaggcc agttttgttc agcggcttgt atgggccagt taaagaatta gaaacataac
     1681 caagcatgta aatatcgtta gacgtaatgc cgtcaatcgt catttttgat ccgcgggagt
     1741 cagtgaacag gtaccatttg ccgttcattt taaagacgtt cgcgcgttca atttcatctg
     1801 ttactgtgtt agatgcaatc agcggtttca tcactttttt cagtgtgtaa tcatcgttta
     1861 gctcaatcat accgagagcg ccgtttgcta actcagccgt gcgtttttta tcgctttgca
     1921 gaagtttttg actttcttga cggaagaatg atgtgctttt gccatagtat gctttgttaa
     1981 ataaagattc ttcgccttgg tagccatctt cagttccagt gtttgcttca aatactaagt
     2041 atttgtggcc tttatcttct acgtagtgag gatctctcag cgtatggttg tcgcctgagc
     2101 tgtagttgcc ttcatcgatg aactgctgta cattttgata cgtttttccg tcaccgtcaa
     2161 agattgattt ataatcctct acaccgttga tgttcaaaga gctgtctgat gctgatacgt
     2221 taacttgtgc agttgtcagt gtttgtttgc cgtaatgttt accggagaaa tcagtgtaga
     2281 ataaacggat ttttccgtca gatgtaaatg tggctgaacc tgaccattct tgtgtttggt
     2341 cttttaggat agaatcattt gcatcgaatt tgtcgctgtc tttaaagacg cggccagcgt
     2401 ttttccagct gtcaatagaa gtttcgccga ctttttgata gaacatgtaa atcgatgtgt
     2461 catccgcatt tttaggatct ccggctaatg caaagacgat gtggtagccg tgatagtttg
     2521 cgacagtgcc gtcagcgttt tgtaatggcc agctgtccca aacgtccagg ccttttgcag
     2581 aagagatatt tttaattgtg gacgaatcga attcaggaac ttgatatttt tcattttttt
     2641 gctgttcagg gatttgcagc atatcatggc gtgtaatatg ggaaatgccg tatgtttcct
     2701 tatatggctt ttggttcgtt tctttcgcaa acgcttgagt tgcgcctcct gccagcagtg
     2761 cggtagtaaa ggttaatact gttgcttgtt ttgcaaactt tttgatgttc atcgttcatg
     2821 tctccttttt tatgtactgt gttagcggtc tgcttcttcc agccctcctg tttgaagatg
     2881 gcaagttagt tacgcacaat aaaaaaagac ctaaaatatg taaggggtga cgccaaagta
     2941 tacactttgc cctttacaca ttttaggtct tgcctgcttt atcagtaaca aacccgcgcg
     3001 atttactttt cgacctcatt ctattagact ctcgtttgga ttgcaactgg tctattttcc
     3061 tcttttgttt gatagaaaat cataaaagga tttgcagact acgggcctaa agaactaaaa
     3121 aatctatctg tttcttttca ttctctgtat tttttatagt ttctgttgca tgggcataaa
     3181 gttgcctttt taatcacaat tcagaaaata tcataatatc tcatttcact aaataatagt
     3241 gaacggcagg tatatgtgat gggttaaaaa ggatcgatcc tctagagtcg acggtatcga
     3301 taagcttgat atcgaattcc tgcagcccgg gggatccact agttctagag cggccgccac
     3361 cgcggccctc tcacttccct gttaagtatc ttcctggcat cttccaggaa atctccgccc
     3421 cgttcgtaag ccatttccgc tcgccgcagt cgaacgaccg agcgtagcga gtcagtgagc
     3481 gaggaagcgg aatatatcct gtatcacata ttctgctgac gcaccggtgc agcctttttt
     3541 ctcctgccac atgaagcgat ccgtccttgg ctgcaggtcg acggatcttt tccgctgcat
     3601 aaccctgctt cggggtcatt atagcgattt tttcggtata tccatccttt ttcgcacgat
     3661 atacaggatt ttgccaaagg gttcgtgtag actttccttg gtgtatccaa cggcgtcagc
     3721 cgggcaggat aggtgaagta ggcccacccg cgagcgggtg ttccttcttc actgtccctt
     3781 attcgcacct ggcggtgctc aacgggaatc ctgctctgcg aggctggccg gctaccgccg
     3841 gcgtaacaga tgagggcaag cggatggctg atgaaaccaa gccaaccagg aagggcagcc
     3901 cacctatcaa ggtgtactgc cttccagacg aacgaagagc gattgaggaa aaggcggcgg
     3961 cggccggcat gagcctgtcg gcctacctgc tggccgtcgg ccagggctac aaaatcacgg
     4021 gcgtcgtgga ctatgagcac gtccgcgagc tggcccgcat caatggcgac ctgggccgcc
     4081 tgggcggcct gctgaaactc tggctcaccg acgacccgcg cacggcgcgg ttcggtgatg
     4141 ccacgatcct cgccctgctg gcgaagatcg aagagaagca ggacgagctt ggcaaggtca
     4201 tgatgggcgt ggtccgcccg agggcagagc catgactttt ttagccgcta aaacggccgg
     4261 ggggtgcgcg tgattgccaa gcacgtcccc atgcgctcca tcaagaagag cgacttcgcg
     4321 gagctggtga agtacatcac cgacgagcaa ggcaagaccg atccccgggg acctgcaggt
     4381 ccggatcact tcactgacac cctcatcagt gccaacatag taagccagta tacactccgc
     4441 tagcgctgat gtccggcggt gcttttgccg ttacgcacca ccccgtcagt agctgaacag
     4501 gagggacagc tgatagaaac agaagccact ggagcacctc aaaaacacca tcatacacta
     4561 aatcagtaag ttggcagcat cacccgacgc actttgcgcc gaataaatac ctgtgacgga
     4621 agatcacttc gcagaataaa taaatcctgg tgtccctgtt gataccggga agccctgggc
     4681 caacttttgg cgaaaatgag acgttgatcg gcacgtaaga ggttccaact ttcaccataa
     4741 tgaaataaga tcactaccgg gcgtattttt tgagttatcg agattttcag gagctaagga
     4801 agctaaaatg gagaaaaaaa tcactggata taccaccgtt gatatatccc aatggcatcg
     4861 taaagaacat tttgaggcat ttcagtcagt tgctcaatgt acctataacc agaccgttca
     4921 gctggatatt acggcctttt taaagaccgt aaagaaaaat aagcacaagt tttatccggc
     4981 ctttattcac attcttgccc gcctgatgaa tgctcatccg gaattccgta tggcaatgaa
     5041 agacggtgag ctggtgatat gggatagtgt tcacccttgt tacaccgttt tccatgagca
     5101 aactgaaacg ttttcatcgc tctggagtga ataccacgac gatttccggc agtttctaca
     5161 catatattcg caagatgtgg cgtgttacgg tgaaaacctg gcctatttcc ctaaagggtt
     5221 tattgagaat atgtttttcg tctcagccaa tccctgggtg agtttcacca gttttgattt
     5281 aaacgtggcc aatatggaca acttcttcgc ccccgttttc accatgggca aatattatac
     5341 gcaaggcgac aaggtgctga tgccgctggc gattcaggtt catcatgccg tttgtgatgg
     5401 cttccatgtc ggcagaatgc ttaatgaatt acaacagtac tgcgatgagt ggcagggcgg
     5461 ggcgtaattt ttttaaggca gttattggtg cccttaaacg cctggttgct acgcctgaat
     5521 aagtgataat aagcggatga atggcagaaa ttcgaaagca aattcgaccc ggtcgtcggt
     5581 tcagggcagg gtcgttaaat agccgcttat gtctattgct ggtttaccgg tttattgact
     5641 accggaagca gtgtgaccgt gtgcttctca aatgcctgag gccagtttgc tcaggctctc
     5701 cccgtggagg taataattga cgatagatcc ggaaccctta atataacttc gtataatgta
     5761 tgctatacga agttattagg tccctcgagg ctgcctcgcg cgtttcggtg atgacggtga
     5821 aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg
     5881 gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gcgcagccat
     5941 gacccagtca cgtagcgata gcggagtgta tactggctta actatgcggc atcagagcag
     6001 attgtactga gagtgcacca tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa
     6061 taccgcatca ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg
     6121 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg
     6181 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag
     6241 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga
     6301 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
     6361 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc
     6421 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg
     6481 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc
     6541 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca
     6601 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag
     6661 ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct
     6721 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc
     6781 accgctggta gcggtggttt ttttgtttgc aa
//
