LOCUS       sequence-248793-        4452 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Template for PCR amplification of XdCrtI.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4452)
  AUTHORS   Milne N, Tramontin LRR, Borodina I
  TITLE     A teaching protocol demonstrating the use of EasyClone and 
            CRISPR/Cas9 for metabolic engineering of Saccharomyces cerevisiae 
            and Yarrowia lipolytica.
  JOURNAL   FEMS Yeast Res. 2019 Sep 26. pii: foz062. doi: 
            10.1093/femsyr/foz062.
  PUBMED    31556952
REFERENCE   2  (bases 1 to 4452)
  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: "FEMS Yeast 
            Res. 2019 Sep 26. pii: foz062. doi: 10.1093/femsyr/foz062."
COMMENT     Sequence Label: pCfB8740
FEATURES             Location/Qualifiers
     source          1..4452
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    105..126
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(141..146,147..164,165..171)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    179..195
                     /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     184..206
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     203..219
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     203..219
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2055..2072)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2055..2071)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2064..2086)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(2280..2299)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2399..2421
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2459..2477)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2545..2649
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2650..2718,2719..3510)
                     /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(2868..2887)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3681..4269
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4170..4189
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4423..4440
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 cgcccaatac gcaaaccgcc tctccccgcg cgttggccga ttcattaatg cagctggcac
       61 gacaggtttc ccgactggaa agcgggcagt gagcgcaacg caattaatgt gagttagctc
      121 actcattagg caccccaggc tttacacttt atgcttccgg ctcgtatgtt gtgtggaatt
      181 gtgagcggat aacaatttca cacaggaaac agctatgacc atgattacgc caagcttgca
      241 tgcctgcagg tcgactctag aggatcccca gtgcaggtat gggaaaagaa caagatcagg
      301 ataaacccac agctatcatc gtgggatgtg gtatcggtgg aatcgccact gccgctcgtc
      361 ttgctaaaga aggtttccag gtcacggtgt tcgagaagaa cgactactcc ggaggtcgat
      421 gctctttaat cgagcgagat ggttatcgat tcgatcaggg gcccagtttg ctgctcttgc
      481 cagatctctt caagcagaca ttcgaagatt tgggggagaa gatggaagat tgggtcgatc
      541 tcatcaagtg tgaacccaac tatgtttgcc acttccacga tgaagagact ttcacttttt
      601 caaccgacat ggcgttgctc aagcgggaag tcgagcgttt tgaaggcaaa gatggatttg
      661 atcggttctt gtcgtttatc caagaagccc acagacatta cgagcttgct gtcgttcacg
      721 tcctgcagaa gaacttccct ggcttcgcag cattcttacg gctacagttc attggccaaa
      781 tcctggctct tcaccccttc gagtctatct ggacaagagt ttgtcgatat ttcaagaccg
      841 acagattacg aagagtcttc tcgtttgcag tgatgtacat gggtcaaagc ccatacagtg
      901 cgcccggaac atattccttg ctccaataca ccgaattgac cgagggcatc tggtatccga
      961 gaggaggctt ttggcaggtt cctaatactc ttcttcagat cgtcaagcgc aacaatccct
     1021 cagccaagtt caatttcaac gctccagttt cccaggttct tctctctcct gccaaggacc
     1081 gagcgactgg tgttcgactt gaatccggcg aggaacatca cgccgatgtt gtgattgtca
     1141 atgctgacct cgtttacgcc tccgagcact tgattcctga cgatgccaga aacaagattg
     1201 gccaactggg tgaagtcaag agaagttggt gggctgactt agttggtgga aagaagctca
     1261 agggaagttg cagtagtttg agcttctact ggagcatgga ccgaatcgtg gacggtctgg
     1321 gcggacacaa tatcttcttg gccgaggact tcaagggatc attcgacaca atcttcgagg
     1381 agttgggtct cccagccgat ccttcctttt acgtgaacgt tccctcgcga atcgatcctt
     1441 ctgccgctcc cgaaggcaaa gatgctatcg tcattcttgt gccgtgtggc catatcgacg
     1501 cttcgaaccc tcaagattac aacaagcttg ttgctcgggc aaggaagttt gtgatccaaa
     1561 cgctttccgc caagcttgga cttcccgact ttgaaaaaat gattgtggca gagaaggttc
     1621 acgatgctcc ctcttgggag aaagaattta acctcaagga cggaagcatc ttgggactgg
     1681 ctcacaactt tatgcaagtt cttggtttca ggccgagcac cagacatccc aagtatgaca
     1741 agttgttctt tgtcggggct tcgactcatc ccggaactgg ggttcccatc gtcttggctg
     1801 gagccaagtt aactgccaac caagttctcg aatcctttga ccgatcccca gctccagatc
     1861 ccaatatgtc actctccgta ccatatggaa aacctctcaa atcaaatgga acgggtatcg
     1921 attctcaggt ccagctgaag ttcatggatt tggagagatg ggtatacctt ttggtgttgt
     1981 tgattggggc cgtgatcgct cgatccgttg gtgttcttgc tttctgaatc gcacggggta
     2041 ccgagctcga attcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt
     2101 acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag
     2161 gcccgcaccg atcgcccttc ccaacagttg cgcagcctga atggcgaatg gcgcctgatg
     2221 cggtattttc tccttacgca tctgtgcggt atttcacacc gcatatggtg cactctcagt
     2281 acaatctgct ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac
     2341 gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc
     2401 gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc
     2461 ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca
     2521 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     2581 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     2641 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
     2701 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
     2761 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
     2821 tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg
     2881 gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
     2941 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
     3001 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
     3061 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
     3121 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
     3181 accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt
     3241 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
     3301 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     3361 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
     3421 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
     3481 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
     3541 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     3601 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     3661 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     3721 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     3781 tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag
     3841 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     3901 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     3961 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     4021 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     4081 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     4141 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     4201 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     4261 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     4321 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt
     4381 gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag
     4441 gaagcggaag ag
//
