LOCUS       sequence-253008-        2643 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Level 0 destination vector for a coding sequence with B - C scars. 
            Clone in with BpiI.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2643)
  AUTHORS   Young EM, Zhao Z, Gielesen BEM, Wu L, Benjamin Gordon D, Roubos JA, 
            Voigt CA
  TITLE     Iterative algorithm-guided design of massive strain libraries, 
            applied to itaconic acid production in yeast.
  JOURNAL   Metab Eng. 2018 Jul;48:33-43. doi: 10.1016/j.ymben.2018.05.002. Epub
            2018 May 9.
  PUBMED    29753070
REFERENCE   2  (bases 1 to 2643)
  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: "Metab 
            Eng."; date: "2018-07"; volume: "48:33-43. doi"; pages: " 
            10.1016/j.ymben.2018.05.002. Epub 2018 May 9"
COMMENT     Sequence Label: pEMY07AB-ORF
FEATURES             Location/Qualifiers
     source          1..2643
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     402..421
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    731..752
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(767..772,773..790,791..797)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    805..821
                     /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     810..832
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     829..845
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     829..845
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             841..1179
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLHACRSILEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISRRIWCRRPHLRP
                     STICSDAA"
     misc_feature    858..914
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(915..932)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(915..931)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(924..946)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(1155..1174)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1274..1296
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1334..1352)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1420..1524
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1525..1593,1594..2385)
                     /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(1743..1762)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(2556..2643,1..501)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt
       61 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
      121 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
      181 gggttgggct caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
      241 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
      301 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
      361 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
      421 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
      481 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
      541 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
      601 attaccgcct ttgagtgagc tgataccgct cggtctcttt taaggtcttc cccgcgcgtt
      661 ggccgattca ttaatgcagc tggcacgaca ggtttcccta ctggaaagcg ggcagtgagc
      721 gcaacgcaat taatgtgagt tagctcactc attaggcacc ccaggcttta cactttatgc
      781 ttccggctcg tatgttgtgt ggaattgtga gcggataaca atttcacaca ggaaacagct
      841 atgaccatga ttacgccaag cttgcatgcc tgcaggtcga ttctagagga tccccgggta
      901 ccgagctcga attcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt
      961 acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag
     1021 gcccgcaccg atcgcccttc ccaacagttg cgcagcctga atggcgaatg gcgcctgatg
     1081 cggtattttc tccttacgca tctgtgcggt atttcacgcc gcatatggtg cagaagacct
     1141 catttgagac ccagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg
     1201 ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa
     1261 gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc
     1321 gcgagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg
     1381 gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     1441 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     1501 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     1561 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     1621 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     1681 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     1741 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     1801 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     1861 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     1921 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     1981 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     2041 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     2101 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     2161 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     2221 tctggagccg gtgagcgtgg ctctcgcggt atcattgcag cactggggcc agatggtaag
     2281 ccctcccgta tcgtagttat ctacacgacg gggagccagg caactatgga tgaacgaaat
     2341 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     2401 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     2461 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     2521 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     2581 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     2641 gag
//
