LOCUS       sequence-7023-ex        5138 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5138)
  AUTHORS   Colwill K, Wells CD, Elder K, Goudreault M, Hersi K, Kulkarni S, 
            Hardy WR, Pawson T, Morin GB
  TITLE     Modification of the Creator recombination system for proteomics 
            applications--improved expression by addition of splice sites.
  JOURNAL   BMC Biotechnol. 2006 Mar 6;6:13. doi: 10.1186/1472-6750-6-13.
  PUBMED    16519801
REFERENCE   2  (bases 1 to 5138)
  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: "BMC 
            Biotechnol."; date: "2006-03-6"; volume: "6:13. doi"; pages: " 
            10.1186/1472-6750-6-13"
COMMENT     Sequence Label: V308 pDNR MCS lacZ alpha - Donor
FEATURES             Location/Qualifiers
     source          1..5138
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    complement(1..34)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    149..170
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(185..190,191..208,209..215)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    223..239
                     /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)."
     primer_bind     228..250
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     247..263
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     247..263
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             259..432
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGE"
     primer_bind     complement(279..295)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(288..310)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     terminator      complement(489..583)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(604..1263)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     1178..1197
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     protein_bind    complement(1292..1325)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        1340..1785
                     /gene="sacR"
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     CDS             join(1786..1872,1873..3207)
                     /codon_start=1
                     /gene="Bacillus subtilis sacB"
                     /product="secreted levansucrase that renders bacterial 
                     growth sensitive to sucrose"
                     /label=SacB
                     /note="negative selection marker"
                     /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
                     ITRHDMLQIPEQQKNEKYQVPEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYH
                     IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
                     SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
                     KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
                     YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
                     IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
                     MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
                     VKDSILEQGQLTVNK"
     promoter        3275..3379
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3380..3448,3449..4240)
                     /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(3598..3617)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4411..4999
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4900..4919
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5116..5132
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 ataacttcgt atagcataca ttatacgaag ttatggcgcg ccggcgccca atacgcaaac
       61 cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg tttcccgact
      121 ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta gctcactcat taggcacccc
      181 aggctttaca ctttatgctt ccggctcgta tgttgtgtga aattgtgagc ggataacaat
      241 ttcacacagg aaacagctat gaccatgatt acggattcac tggccgtcgt tttacaacgt
      301 cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc
      361 gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc
      421 ctgaatggcg aatggcgtta attaaccagg taagttccta ggagatcctg gtcatgacta
      481 gtgcttggat tctcaccaat aaaaaacgcc cggcggcaac cgagcgttct gaacaaatcc
      541 agatggagtt ctgaggtcat tactggatct atcaacagga gtccaagcga gctcgatatc
      601 aaattacgcc ccgccctgcc actcatcgca gtactgttgt aattcattaa gcattctgcc
      661 gacatggaag ccatcacaaa cggcatgatg aacctgaatc gccagcggca tcagcacctt
      721 gtcgccttgc gtataatatt tgcccatggt gaaaacgggg gcgaagaagt tgtccatatt
      781 ggccacgttt aaatcaaaac tggtgaaact cacccaggga ttggctgaga cgaaaaacat
      841 attctcaata aaccctttag ggaaataggc caggttttca ccgtaacacg ccacatcttg
      901 cgaatatatg tgtagaaact gccggaaatc gtcgtggtat tcactccaga gcgatgaaaa
      961 cgtttcagtt tgctcatgga aaacggtgta acaagggtga acactatccc atatcaccag
     1021 ctcaccgtct ttcattgcca tacgaaattc cggatgagca ttcatcaggc gggcaagaat
     1081 gtgaataaag gccggataaa acttgtgctt atttttcttt acggtcttta aaaaggccgt
     1141 aatatccagc tgaacggtct ggttataggt acattgagca actgactgaa atgcctcaaa
     1201 atgttcttta cgatgccatt gggatatatc aacggtggta tatccagtga tttttttctc
     1261 cattttagct tccttagctc ctgaaagatc cataacttcg tatagcatac attatacgaa
     1321 gttatgcggc cgcgacgtcc acatatacct gccgttcact attatttagt gaaatgagat
     1381 attatgatat tttctgaatt gtgattaaaa aggcaacttt atgcccatgc aacagaaact
     1441 ataaaaaata cagagaatga aaagaaacag atagattttt tagttcttta ggcccgtagt
     1501 ctgcaaatcc ttttatgatt ttctatcaaa caaaagagga aaatagacca gttgcaatcc
     1561 aaacgagagt ctaatagaat gaggtcgaaa agtaaatcgc gcgggtttgt tactgataaa
     1621 gcaggcaaga cctaaaatgt gtaaagggca aagtgtatac tttggcgtca ccccttacat
     1681 attttaggtc tttttttatt gtgcgtaact aacttgccat cttcaaacag gagggctgga
     1741 agaagcagac cgctaacaca gtacataaaa aaggagacat gaacgatgaa catcaaaaag
     1801 tttgcaaaac aagcaacagt attaaccttt actaccgcac tgctggcagg aggcgcaact
     1861 caagcgtttg cgaaagaaac gaaccaaaag ccatataagg aaacatacgg catttcccat
     1921 attacacgcc atgatatgct gcaaatccct gaacagcaaa aaaatgaaaa atatcaagtt
     1981 cctgagttcg attcgtccac aattaaaaat atctcttctg caaaaggcct ggacgtttgg
     2041 gacagctggc cattacaaaa cgctgacggc actgtcgcaa actatcacgg ctaccacatc
     2101 gtctttgcat tagccggaga tcctaaaaat gcggatgaca catcgattta catgttctat
     2161 caaaaagtcg gcgaaacttc tattgacagc tggaaaaacg ctggccgcgt ctttaaagac
     2221 agcgacaaat tcgatgcaaa tgattctatc ctaaaagacc aaacacaaga atggtcaggt
     2281 tcagccacat ttacatctga cggaaaaatc cgtttattct acactgattt ctccggtaaa
     2341 cattacggca aacaaacact gacaactgca caagttaacg tatcagcatc agacagctct
     2401 ttgaacatca acggtgtaga ggattataaa tcaatctttg acggtgacgg aaaaacgtat
     2461 caaaatgtac agcagttcat cgatgaaggc aactacagct caggcgacaa ccatacgctg
     2521 agagatcctc actacgtaga agataaaggc cacaaatact tagtatttga agcaaacact
     2581 ggaactgaag atggctacca aggcgaagaa tctttattta acaaagcata ctatggcaaa
     2641 agcacatcat tcttccgtca agaaagtcaa aaacttctgc aaagcgataa aaaacgcacg
     2701 gctgagttag caaacggcgc tctcggtatg attgagctaa acgatgatta cacactgaaa
     2761 aaagtgatga aaccgctgat tgcatctaac acagtaacag atgaaattga acgcgcgaac
     2821 gtctttaaaa tgaacggcaa atggtacctg ttcactgact cccgcggatc aaaaatgacg
     2881 attgacggca ttacgtctaa cgatatttac atgcttggtt atgtttctaa ttctttaact
     2941 ggcccataca agccgctgaa caaaactggc cttgtgttaa aaatggatct tgatcctaac
     3001 gatgtaacct ttacttactc acacttcgct gtacctcaag cgaaaggaaa caatgtcgtg
     3061 attacaagct atatgacaaa cagaggattc tacgcagaca aacaatcaac gtttgcgcct
     3121 agcttcctgc tgaacatcaa aggcaagaaa acatctgttg tcaaagacag catccttgaa
     3181 caaggacaat taacagttaa caaataaaaa cgcaaaagaa aatgccgata tcctattggc
     3241 attgacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     3301 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     3361 atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt
     3421 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc
     3481 tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat
     3541 ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct
     3601 atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca
     3661 ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg
     3721 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa
     3781 cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg
     3841 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga
     3901 cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg
     3961 cgaactactt actctagctt cccggcaaca attaatagac tggatggagg cggataaagt
     4021 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg
     4081 agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc
     4141 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     4201 gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc
     4261 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     4321 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     4381 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     4441 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     4501 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct
     4561 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     4621 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     4681 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     4741 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     4801 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     4861 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     4921 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     4981 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     5041 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
     5101 taccgcctta cgcgtgtaaa acgacggcca gtagatct
//
