LOCUS       sequence-12876-e        7153 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 7153)
  AUTHORS   Cross FR, Pecani K
  TITLE     Efficient and rapid exact gene replacement without selection.
  JOURNAL   Yeast. 2011 Feb . 28(2):167-79.
  PUBMED    21246629
REFERENCE   2  (bases 1 to 7153)
  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: "Yeast. 2011
            Feb . 28(2):167-79."
COMMENT     Sequence Label: FC858
FEATURES             Location/Qualifiers
     source          1..7153
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(10..28)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        96..200
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(201..269,270..1061)
                     /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(419..438)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1232..1820
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1721..1740
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1974..1991
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        join(2144..2149,2150..2167,2168..2174)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2182..2198
                     /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     2187..2209
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2206..2222
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2206..2222
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     2241..2261
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        2243..2261
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5117..5133)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5118..5134)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     complement(5158..5177)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5159..5177)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5187..5204)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5187..5203)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5196..5218)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5348..5803
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5435..5454)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5645..5666
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(5934..6737)
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     promoter        complement(6738..6958)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(6984..7003)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7103..7125
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
      121 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
      181 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
      241 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
      301 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
      361 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
      421 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
      481 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
      541 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
      601 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
      661 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
      721 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
      781 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
      841 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
      901 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
      961 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     1021 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     1081 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     1141 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     1201 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     1261 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     1321 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc
     1381 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     1441 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     1501 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     1561 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     1621 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     1681 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     1741 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     1801 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     1861 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     1921 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     1981 cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc
     2041 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca
     2101 acgcaattaa tgtgagttac ctcactcatt aggcacccca ggctttacac tttatgcttc
     2161 cggctcctat gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg
     2221 accatgatta cgccaagcgc gcaattaacc ctcactaaag ggaacaaaag ctggagctcc
     2281 accgcggtgg cggccgcaac atctcagtta cccgcactgc cttttttctc tcccgctttt
     2341 cagcagaacg aggcgacaag cgaaggaaca aaacaaaaca aaagaaaaca aaaatatccc
     2401 gggaatctcg gtcgtaatga tttttataat gacgaaaaaa aaaaaattgg aaagaaaaag
     2461 ctttaatgcg gtagtttatc acagttaaat tgctaacgca gtcaggcacc gtgtatgaaa
     2521 tctaacaatg cgctcatcgt catcctcggc accgtcaccc tggatgctgt aggcataggc
     2581 ttggttatgc cggtactgcc gggcctcttg cgggatatcg tccattccga cagcatcgcc
     2641 agtcactatg gcgtgctgct agcgctatat gcgttgatgc aatttctatg cgcacccgtt
     2701 ctcggagcac tgtccgaccg ctttggccgc cgcccagtcc tgctcgcttc gctacttgga
     2761 gccactatcg actacgcgat catggcgacc acacccgtcc tgtggatctt atattctcca
     2821 gattccaaag ctagaaagct gaagtcaccg gaaggagagc cctgttggtg gactacccta
     2881 tgattattat caaagacttc ttccacatct atgacaatgc catagtcatt ataactggga
     2941 agcgcataat ctgccaactt atcatcgtaa acttccaaat tgtaagaacc ttttaaaagg
     3001 gtgtctttgg atagttcctt caggaagcat tttctgtctc caccataacc atagaagtaa
     3061 aacgcggaaa ctctgaatgg caataaaacg aatgccagga agatgtagtg tgataacatt
     3121 ggcaaatcta gatcctttca gattgaggcg ataaggaaga aagcgccaac aatctgttca
     3181 caacctcacc aataaagtct atcagtagag attttaacta tacaccccat atcttgttgt
     3241 gtcgatattg gtttcttacc cgaagaacga gtttctaatt ccacgacgtt tttttatccc
     3301 acacaaaatg gttttttgca ccgcttctag cattaatatt gcgatgaaga agaacctcat
     3361 ggatttacaa taccttagac ttggccatct caaacttggt tcatcctcta actccgattt
     3421 ttgtcaccct tccttctagt cttcatccta aatttagttg catgtccaca aacctgaacc
     3481 cattcatgaa taatgcgcct tccagtgcac cactcaaggg ttctgaaagt aagagggtat
     3541 cgaaaaggcc aatatctagt tcttcgtccg cctcactatt atcagctccc tccaggcgat
     3601 cgaggccgtc tacagtatat ggtgataggt atattccttc ccgaacagat atagatttca
     3661 attccatcgt ttccatcagt agtatggcaa gcgttccagc ccttaatcca tcgagtacgg
     3721 aagatcaagt agaataccaa aaggaaagac aggcacatga aacttataat actttgctga
     3781 agaatgagct ttttggcgag atgctaagca aggatacggt aggctcagag agcagtatag
     3841 atcgcattaa gaatacgaga ccatctacta gaggcaatgt gcacgcagaa aatacaacta
     3901 gacatggtta tgaactggaa cgagtaagcg ctccacctcc tgaagctgct ggtttagagg
     3961 aatttgcgcc acatagtgca ccagtcgctc ctcgacgtct ttttacatca cagcaagatg
     4021 aaatcacgag accctcaagc aactcagtta ggggcgccag cctgttgact tatcaacaac
     4081 gtaaaggtag aagactatct gcggcctctt tgttacaatc tcaatttttt gattccatgg
     4141 cgccagttag gccggattcc aaacagctcc tgctagcacc ggggaagcaa tttagacaaa
     4201 tagctaaagt tccctataga gttttagatg ccccatcatt agcagacgat ttttactaca
     4261 gcttgataga ttggtcaagt actgatgttt tggcagttgc tcttggaaaa tcgatttttt
     4321 taaccgataa taacactggc gacgttgttc acttatgcga cacggaaaac gaatacacaa
     4381 gcttaagctg gattggagca ggctctcatc tggcagtagg tcaagcaaat ggacttgtag
     4441 aaatttatga cgtaatgaaa agaaaatgta ttaggacgtt gtcgggacat attgacagag
     4501 tagcgtgttt atcttggaat aatcatgttt tgacgtctgg gagtagagat catcgaatac
     4561 tgcataggga tgttaggatg ccagatccct tttttgaaac tatagaatcg cacactcagg
     4621 aagtctgtgg cctaaagtgg aatgtagcgg acaacaaact tgcctcaggt ggtaacgata
     4681 acgtcgtcca tgtttacgaa ggaacgtcga aagctccaat cttgacgttt gacgaacata
     4741 aggctgctgt taaggcaatg gcctgggccc ctcataaacg aggtgtgcta gctaccggtg
     4801 gtggtacagc agatagaaga ctaaaaatct ggaacgttaa tacgtcaata aagatgagtg
     4861 atatagatag cggctctcaa atatgtaata tggtatggtc aaagaatact aacgagcttg
     4921 tgacatcaca tggttactca aaatataact taaccttgtg ggactgtaac tctatggatc
     4981 taaataaatt cgttttcaat gattaaaata gcatagtcgg gtttttcttt tagtttcagc
     5041 tttccgcaac agtataattt tataaaccct ggttttggtt ttgtagagtg gttgacgaat
     5101 tcgatatcaa gcttatcgat accgtcgacc tcgagggggg gcccggtacc caattcgccc
     5161 tatagtgagt cgtattacgc gcgctcactg gccgtcgttt tacaacgtcg tgactgggaa
     5221 aaccctggcg ttacccaact taatcgcctt gcagcacatc cccctttcgc cagctggcgt
     5281 aatagcgaag aggcccgcac cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa
     5341 tggcgcgacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     5401 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     5461 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     5521 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     5581 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     5641 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     5701 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     5761 aaatttaacg cgaattttaa caaaatatta acgtttacaa tttcctgatg cggtattttc
     5821 tccttacgca tctgtgcggt atttcacacc gcatagggta ataactgata taattaaatt
     5881 gaagctctaa tttgtgagtt tagtatacat gcatttactt ataatacagt tttttagttt
     5941 tgctggccgc atcttctcaa atatgcttcc cagcctgctt ttctgtaacg ttcaccctct
     6001 accttagcat cccttccctt tgcaaatagt cctcttccaa caataataat gtcagatcct
     6061 gtagagacca catcatccac ggttctatac tgttgaccca atgcgtctcc cttgtcatct
     6121 aaacccacac cgggtgtcat aatcaaccaa tcgtaacctt catctcttcc acccatgtct
     6181 ctttgagcaa taaagccgat aacaaaatct ttgtcgctct tcgcaatgtc aacagtaccc
     6241 ttagtatatt ctccagtaga tagggagccc ttgcatgaca attctgctaa catcaaaagg
     6301 cctctaggtt cctttgttac ttcttctgcc gcctgcttca aaccgctaac aatacctggg
     6361 cccaccacac cgtgtgcatt cgtaatgtct gcccattctg ctattctgta tacacccgca
     6421 gagtactgca atttgactgt attaccaatg tcagcaaatt ttctgtcttc gaagagtaaa
     6481 aaattgtact tggcggataa tgcctttagc ggcttaactg tgccctccat ggaaaaatca
     6541 gtcaagatat ccacatgtgt ttttagtaaa caaattttgg gacctaatgc ttcaactaac
     6601 tccagtaatt ccttggtggt acgaacatcc aatgaagcac acaagtttgt ttgcttttcg
     6661 tgcatgatat taaatagctt ggcagcaaca ggactaggat gagtagcagc acgttcctta
     6721 tatgtagctt tcgacatgat ttatcttcgt ttcctgcagg tttttgttct gtgcagttgg
     6781 gttaagaata ctgggcaatt tcatgtttct tcaacactac atatgcgtat atataccaat
     6841 ctaagtctgt gctccttcct tcgttcttcc ttctgttcgg agattaccga atcaaaaaaa
     6901 tttcaaagaa accgaaatca aaaaaaagaa taaaaaaaaa atgatgaatt gaattgaaaa
     6961 gctgtggtat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagccc
     7021 cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct
     7081 tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca
     7141 ccgaaacgcg cga
//
