LOCUS       sequence-7025-ex        4788 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 4788)
  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 4788)
  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: V624 pDNR MCS BE - Donor
FEATURES             Location/Qualifiers
     source          1..4788
                     /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)."
     terminator      complement(111..205)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(226..885)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     800..819
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     protein_bind    complement(914..947)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        962..1407
                     /gene="sacR"
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     CDS             join(1408..1494,1495..2829)
                     /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        2897..3001
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3002..3070,3071..3862)
                     /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(3220..3239)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4033..4621
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4522..4541
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4737..4754
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4738..4754
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4762..4781
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4762..4780
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ataacttcgt atagcataca ttatacgaag ttatggggat ccgtcgaccc cgggaagctt
       61 gaattccagg taagttagcc taggagatcc tggtcatgac tagtgcttgg attctcacca
      121 ataaaaaacg cccggcggca accgagcgtt ctgaacaaat ccagatggag ttctgaggtc
      181 attactggat ctatcaacag gagtccaagc gagctcgata tcaaattacg ccccgccctg
      241 ccactcatcg cagtactgtt gtaattcatt aagcattctg ccgacatgga agccatcaca
      301 aacggcatga tgaacctgaa tcgccagcgg catcagcacc ttgtcgcctt gcgtataata
      361 tttgcccatg gtgaaaacgg gggcgaagaa gttgtccata ttggccacgt ttaaatcaaa
      421 actggtgaaa ctcacccagg gattggctga gacgaaaaac atattctcaa taaacccttt
      481 agggaaatag gccaggtttt caccgtaaca cgccacatct tgcgaatata tgtgtagaaa
      541 ctgccggaaa tcgtcgtggt attcactcca gagcgatgaa aacgtttcag tttgctcatg
      601 gaaaacggtg taacaagggt gaacactatc ccatatcacc agctcaccgt ctttcattgc
      661 catacgaaat tccggatgag cattcatcag gcgggcaaga atgtgaataa aggccggata
      721 aaacttgtgc ttatttttct ttacggtctt taaaaaggcc gtaatatcca gctgaacggt
      781 ctggttatag gtacattgag caactgactg aaatgcctca aaatgttctt tacgatgcca
      841 ttgggatata tcaacggtgg tatatccagt gatttttttc tccattttag cttccttagc
      901 tcctgaaaga tccataactt cgtatagcat acattatacg aagttatgcg gccgcgacgt
      961 ccacatatac ctgccgttca ctattattta gtgaaatgag atattatgat attttctgaa
     1021 ttgtgattaa aaaggcaact ttatgcccat gcaacagaaa ctataaaaaa tacagagaat
     1081 gaaaagaaac agatagattt tttagttctt taggcccgta gtctgcaaat ccttttatga
     1141 ttttctatca aacaaaagag gaaaatagac cagttgcaat ccaaacgaga gtctaataga
     1201 atgaggtcga aaagtaaatc gcgcgggttt gttactgata aagcaggcaa gacctaaaat
     1261 gtgtaaaggg caaagtgtat actttggcgt caccccttac atattttagg tcttttttta
     1321 ttgtgcgtaa ctaacttgcc atcttcaaac aggagggctg gaagaagcag accgctaaca
     1381 cagtacataa aaaaggagac atgaacgatg aacatcaaaa agtttgcaaa acaagcaaca
     1441 gtattaacct ttactaccgc actgctggca ggaggcgcaa ctcaagcgtt tgcgaaagaa
     1501 acgaaccaaa agccatataa ggaaacatac ggcatttccc atattacacg ccatgatatg
     1561 ctgcaaatcc ctgaacagca aaaaaatgaa aaatatcaag ttcctgagtt cgattcgtcc
     1621 acaattaaaa atatctcttc tgcaaaaggc ctggacgttt gggacagctg gccattacaa
     1681 aacgctgacg gcactgtcgc aaactatcac ggctaccaca tcgtctttgc attagccgga
     1741 gatcctaaaa atgcggatga cacatcgatt tacatgttct atcaaaaagt cggcgaaact
     1801 tctattgaca gctggaaaaa cgctggccgc gtctttaaag acagcgacaa attcgatgca
     1861 aatgattcta tcctaaaaga ccaaacacaa gaatggtcag gttcagccac atttacatct
     1921 gacggaaaaa tccgtttatt ctacactgat ttctccggta aacattacgg caaacaaaca
     1981 ctgacaactg cacaagttaa cgtatcagca tcagacagct ctttgaacat caacggtgta
     2041 gaggattata aatcaatctt tgacggtgac ggaaaaacgt atcaaaatgt acagcagttc
     2101 atcgatgaag gcaactacag ctcaggcgac aaccatacgc tgagagatcc tcactacgta
     2161 gaagataaag gccacaaata cttagtattt gaagcaaaca ctggaactga agatggctac
     2221 caaggcgaag aatctttatt taacaaagca tactatggca aaagcacatc attcttccgt
     2281 caagaaagtc aaaaacttct gcaaagcgat aaaaaacgca cggctgagtt agcaaacggc
     2341 gctctcggta tgattgagct aaacgatgat tacacactga aaaaagtgat gaaaccgctg
     2401 attgcatcta acacagtaac agatgaaatt gaacgcgcga acgtctttaa aatgaacggc
     2461 aaatggtacc tgttcactga ctcccgcgga tcaaaaatga cgattgacgg cattacgtct
     2521 aacgatattt acatgcttgg ttatgtttct aattctttaa ctggcccata caagccgctg
     2581 aacaaaactg gccttgtgtt aaaaatggat cttgatccta acgatgtaac ctttacttac
     2641 tcacacttcg ctgtacctca agcgaaagga aacaatgtcg tgattacaag ctatatgaca
     2701 aacagaggat tctacgcaga caaacaatca acgtttgcgc ctagcttcct gctgaacatc
     2761 aaaggcaaga aaacatctgt tgtcaaagac agcatccttg aacaaggaca attaacagtt
     2821 aacaaataaa aacgcaaaag aaaatgccga tatcctattg gcattgacgt caggtggcac
     2881 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
     2941 gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag
     3001 tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc
     3061 tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc
     3121 acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc
     3181 cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc
     3241 ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt
     3301 ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt
     3361 atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat
     3421 cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct
     3481 tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat
     3541 gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc
     3601 ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg
     3661 ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc
     3721 tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta
     3781 cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc
     3841 ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga
     3901 tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat
     3961 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
     4021 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     4081 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     4141 ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt
     4201 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     4261 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     4321 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     4381 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     4441 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     4501 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     4561 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
     4621 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
     4681 gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct tacgcgtgta
     4741 aaacgacggc cagtagatct gtaatacgac tcactatagg gcgctagc
//
