LOCUS       sequence-462975-        4205 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Vector part for BEVA; BEVA2.0 Position 4 sacB sucrose 
            counterselection module, AmpR..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4205)
  AUTHORS   Geddes BA, Williamson R, Schumacher J, Ardi A, Levin G, Cervenka E, 
            Huang R, diCenzo G
  TITLE     BEVA2.0: modular assembly of golden gate-compatible vectors with 
            expanded utility for genetic engineering.
  JOURNAL   Can J Microbiol. 2025 Jan 1;71:1-13. doi: 10.1139/cjm-2024-0246.
  PUBMED    39919271
REFERENCE   2  (bases 1 to 4205)
  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: "Can J 
            Microbiol."; date: "2025-01-1"; volume: "71:1-13. doi"; pages: " 
            10.1139/cjm-2024-0246"
COMMENT     Sequence Label: pNDGG008
FEATURES             Location/Qualifiers
     source          1..4205
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        21..466
                     /gene="sacR"
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     CDS             join(467..553,554..1888)
                     /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
                     ITRHDMLQIPEQQKNEKYQVSEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYH
                     IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
                     SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
                     KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
                     YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
                     IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
                     MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
                     VKDSILEQGQLTVNK"
     primer_bind     complement(2041..2057)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2041..2057)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2054..2076)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     rep_origin      complement(2229..2817)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2309..2328)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    complement(2934..2958)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(join(3058..3849,3850..3918))
                     /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     3681..3700
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3919..4023)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4122..4144
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4136..4153
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4137..4153
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 cactctgtgc gtctcattac cacatatacc tgccgttcac tattatttag tgaaatgaga
       61 tattatgata ttttctgaat tgtgattaaa aaggcaactt tatgcccatg caacagaaac
      121 tataaaaaat acagagaatg aaaagaaaca gatagatttt ttagttcttt aggcccgtag
      181 tctgcaaatc cttttatgat tttctatcaa acaaaagagg aaaatagacc agttgcaatc
      241 caaacgagag tctaatagaa tgaggtcgaa aagtaaatcg cgcgggtttg ttactgataa
      301 agcaggcaag acctaaaatg tgtaaagggc aaagtgtata ctttggcgtc accccttaca
      361 tattttaggt ctttttttat tgtgcgtaac taacttgcca tcttcaaaca ggagggctgg
      421 aagaagcaga ccgctaacac agtacataaa aaaggagaca tgaacgatga acatcaaaaa
      481 gtttgcaaaa caagcaacag tattaacctt tactaccgca ctgctggcag gaggcgcaac
      541 tcaagcgttt gcgaaagaaa cgaaccaaaa gccatataag gaaacatacg gcatttccca
      601 tattacacgc catgatatgc tgcaaatccc tgaacagcaa aaaaatgaaa aatatcaagt
      661 ttctgaattt gattcgtcca caattaaaaa tatctcttct gcaaaaggcc tggacgtttg
      721 ggacagctgg ccattacaaa acgctgacgg cactgtcgca aactatcacg gctaccacat
      781 cgtctttgca ttagccggag atcctaaaaa tgcggatgac acatcgattt acatgttcta
      841 tcaaaaagtc ggcgaaactt ctattgacag ctggaaaaac gctggccgcg tctttaaaga
      901 cagcgacaaa ttcgatgcaa atgattctat cctaaaagac caaacacaag aatggtcagg
      961 ttcagccaca tttacatctg acggaaaaat ccgtttattc tacactgatt tctccggtaa
     1021 acattacggc aaacaaacac tgacaactgc acaagttaac gtatcagcat cagacagctc
     1081 tttgaacatc aacggtgtag aggattataa atcaatcttt gacggtgacg gaaaaacgta
     1141 tcaaaatgta cagcagttca tcgatgaagg caactacagc tcaggcgaca accatacgct
     1201 gagagatcct cactacgtag aagataaagg ccacaaatac ttagtatttg aagcaaacac
     1261 tggaactgaa gatggctacc aaggcgaaga atctttattt aacaaagcat actatggcaa
     1321 aagcacatca ttcttccgtc aagaaagtca aaaacttctg caaagcgata aaaaacgcac
     1381 ggctgagtta gcaaacggcg ctctcggtat gattgagcta aacgatgatt acacactgaa
     1441 aaaagtgatg aaaccgctga ttgcatctaa cacagtaaca gatgaaattg aacgcgcgaa
     1501 cgtctttaaa atgaacggca aatggtatct gttcactgac tcccgcggat caaaaatgac
     1561 gattgacggc attacgtcta acgatattta catgcttggt tatgtttcta attctttaac
     1621 tggcccatac aagccgctga acaaaactgg ccttgtgtta aaaatggatc ttgatcctaa
     1681 cgatgtaacc tttacttact cacacttcgc tgtacctcaa gcgaaaggaa acaatgtcgt
     1741 gattacaagc tatatgacaa acagaggatt ctacgcagac aaacaatcaa cgtttgcgcc
     1801 gagcttcctg ctgaacatca aaggcaagaa aacatctgtt gtcaaagaca gcatccttga
     1861 acaaggacaa ttaacagtta ataaataaaa acgcaaaaga aatgccgatg ggtacgtacc
     1921 cgctcttgga ctcctgttac tccggatccg aattcggcag gcatcagatg agacgcacga
     1981 agtgaggcta ggtggaggct cagtgatgat aagtctgcga tggtggatgc atgtgtcatg
     2041 gtcatagctg tttcctgtgt gaaattgtta tccgctcaga gggcacaatc ctattccgcg
     2101 ctatccgaca atctccaaga cattaggtgg agttcagttc ggcgtatggc atatgtcgct
     2161 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     2221 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     2281 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     2341 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     2401 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     2461 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     2521 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     2581 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     2641 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
     2701 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     2761 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     2821 gatcctttga tcttttctac ggggtctgac gctctattca acaaagccgc cgtcccgtca
     2881 agtcagcgta aatgggtagg gggcttcaaa tcgtcctcgt gataccaatt cggagcctgc
     2941 ttttttgtac aaacttgttg ataatggcaa ttcaaggatc ttcacctaga tccttttaaa
     3001 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     3061 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     3121 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     3181 tgctgcaatg ataccgcgag agccacgctc accggctcca gatttatcag caataaacca
     3241 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     3301 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     3361 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     3421 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     3481 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     3541 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     3601 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     3661 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     3721 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     3781 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     3841 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     3901 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     3961 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     4021 gcgcacattt ccccgaaaag tgccagatac ctgaaacaaa acccatcgta cggccaagga
     4081 agtctccaat aactgtgatc caccacaagc gccagggttt tcccagtcac gacgttgtaa
     4141 aacgacggcc agtcatgcat aatccgcacg catctggaat aaggaagtgc cattccgcct
     4201 gacct
//
