LOCUS       sequence-83388-e        6514 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 6514)
  AUTHORS   Ito-Harashima S, McCusker JH
  TITLE     Positive and negative selection LYS5MX gene replacement cassettes 
            for use in Saccharomyces cerevisiae.
  JOURNAL   Yeast. 2004 Jan 15;21(1):53-61.
  PUBMED    14745782
REFERENCE   2  (bases 1 to 6514)
  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."; 
            date: "2004-01-15"; volume: "21(1)"; pages: "53-61"
COMMENT     Sequence Label: pSA49
FEATURES             Location/Qualifiers
     source          1..6514
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        187..467
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     CDS             468..1142
                     /codon_start=1
                     /gene="S. cerevisiae TRP1"
                     /product="phosphoribosylanthranilate isomerase, required 
                     for tryptophan biosynthesis"
                     /label=TRP1
                     /note="yeast auxotrophic marker"
                     /translation="MSVINFTGSSGPLVKVCGLQSTEAAECALDSDADLLGIICVPNRK
                     RTIDPVIARKISSLVKAYKNSSGTPKYLVGVFRNQPKEDVLALVNDYGIDIVQLHGDES
                     WQEYQEFLGLPVIKRLVFPKDCNILLSAASQKPHSFIPLFDSEAGGTGELLDWNSISDW
                     VGRQESPESLHFMLAGGLTPENVGDALRLNGVIGVDVSGGVETNGVKDSNKIANFVKNA
                     KK"
     rep_origin      complement(1239..1694)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1376..1397)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1588..1607
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1824..1846
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1838..1855
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1839..1855
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1862..1881
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1862..1880
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        2015..2358
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     terminator      3354..3551
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     3644..3660
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3644..3660
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(3694..3710)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3695..3711)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     promoter        complement(3740..3758)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3779..3795)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3779..3795)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3792..3814)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3803..3819
                     /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)."
     promoter        complement(join(3827..3833,3834..3851,3852..3857))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(4010..4027)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4181..4769)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(4940..5731,5732..5800))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFFHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5563..5582
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5801..5905)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    5942..6445
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     6487..6505
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accataaacg acattactat atatataata taggaagcat ttaatagaca gcatcgtaat
      241 atatgtgtac tttgcagtta tgacgccaga tggcagtagt ggaagatatt ctttattgaa
      301 aaatagcttg tcaccttacg tacaatcttg atccggagct tttctttttt tgccgattaa
      361 gaattaattc ggtcgaaaaa agaaaaggag agggccaaga gggagggcat tggtgactat
      421 tgagcacgtg agtatacgtg attaagcaca caaaggcagc ttggagtatg tctgttatta
      481 atttcacagg tagttctggt ccattggtga aagtttgcgg cttgcagagc acagaggccg
      541 cagaatgtgc tctagattcc gatgctgact tgctgggtat tatatgtgtg cccaatagaa
      601 agagaacaat tgacccggtt attgcaagga aaatttcaag tcttgtaaaa gcatataaaa
      661 atagttcagg cactccgaaa tacttggttg gcgtgtttcg taatcaacct aaggaggatg
      721 ttttggctct ggtcaatgat tacggcattg atatcgtcca actgcatgga gatgagtcgt
      781 ggcaagaata ccaagagttc ctcggtttgc cagttattaa aagactcgta tttccaaaag
      841 actgcaacat actactcagt gcagcttcac agaaacctca ttcgtttatt cccttgtttg
      901 attcagaagc aggtgggaca ggtgaacttt tggattggaa ctcgatttct gactgggttg
      961 gaaggcaaga gagccccgaa agcttacatt ttatgttagc tggtggactg acgccagaaa
     1021 atgttggtga tgcgcttaga ttaaatggcg ttattggtgt tgatgtaagc ggaggtgtgg
     1081 agacaaatgg tgtaaaagac tctaacaaaa tagcaaattt cgtcaaaaat gctaagaaat
     1141 aggttattac tgagtagtat ttatttaagt attgtttgtg cacttgcctg cggtgtgaaa
     1201 taccgcacag atgcgtaagg agaaaatacc gcatcaggaa attgtaaacg ttaatatttt
     1261 gttaaaattc gcgttaaatt tttgttaaat cagctcattt tttaaccaat aggccgaaat
     1321 cggcaaaatc ccttataaat caaaagaata gaccgagata gggttgagtg ttgttccagt
     1381 ttggaacaag agtccactat taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt
     1441 ctatcagggc gatggcccac tacgtgaacc atcaccctaa tcaagttttt tggggtcgag
     1501 gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc cgatttagag cttgacgggg
     1561 aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc
     1621 gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc
     1681 gctacagggc gcgtcgcgcc attcgccatt caggctgcgc aactgttggg aagggcgatc
     1741 ggtgcgggcc tcttcgctat tacgccagct ggcgaagggg ggatgtgctg caaggcgatt
     1801 aagttgggta acgccagggt tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt
     1861 gtaatacgac tcactatagg gcgaattgga gctccaccgc ggtggcggcc gccagctgaa
     1921 gcttcgtacg ctgcaggtcg acggatcccc gggttaatta aggcgcgcca gatctgttta
     1981 gcttgccttg tccccgccgg gtcacccggc cagcgacatg gaggcccaga ataccctcct
     2041 tgacagtctt gacgtgcgca gctcaggggc atgatgtgac tgtcgcccgt acatttagcc
     2101 catacatccc catgtataat catttgcatc catacatttt gatggccgca cggcgcgaag
     2161 caaaaattac ggctcctcgc tgcagacctg cgagcaggga aacgctcccc tcacagacgc
     2221 gttgaattgt ccccacgccg cgcccctgta gagaaatata aaaggttagg atttgccact
     2281 gaggttcttc tttcatatac ttccttttaa aatcttgcta ggatacagtt ctcacatcac
     2341 atccgaacat aaacaaccat gacattacaa gatcatctta atgataaatc aaatatcatc
     2401 ctacttttca ccactagatt aactaatgaa ttaattgaat tttggaatga tgattataat
     2461 tttgaatgtt ctttaagatt attaaatgat ttatcattac aacaaaaaat atcttcaatt
     2521 acaaaaaacc ccaaccatca acaacaaaga tttaaatata aacaattatt atcaaattta
     2581 tttattagat atgcaatcaa ttcaatcatt caatcaaatg atctatttaa aaaaattgaa
     2641 tttgattata atgaatttgg gaaacccata ttaccatata atttccaatt caatattagt
     2701 agttctaatg atataattgc cttaattgtg gaatttccat cttcatcttc attatcatca
     2761 tcatcattgt catcatcatt atctggtgat cgagcttatg acccaattgg tattgattta
     2821 tcacattcta ttcaaaatag tatatcaaat acagaatatc ttgaacaatt taaaccgatt
     2881 tttgatgatg tttatgaatt accacaaatt gataattatt ttaaatttaa tcatttttgg
     2941 acattaaaag aatcatttac aaaattgatt ggtagtggat taaatattga attatcagat
     3001 ttttatttca tagttgatta taatgataat gagttttcta gtgataatga aaaattccaa
     3061 acaagtatag ttggaaataa tattggagat agttggatgt attattcatt gaattggttt
     3121 gataaaatct tgataaatat agataaactt gaattagcca agaatcaatt tataacggca
     3181 ttaaataaaa aagaatttta ttgtcgtagt tctattcttg ataatggtaa tgtcgatgaa
     3241 tctaaattgc cagtaattat aacgataatt aatcaaaaca agaataaatc aattcaacca
     3301 atagagttga gatttgaaaa attattatct gaacatttaa accagtgatc agtactgaca
     3361 ataaaaagat tcttgttttc aagaacttgt catttgtata gtttttttat attgtagttg
     3421 ttctatttta atcaaatgtt agcgtgattt atattttttt tcgcctcgac atcatctgcc
     3481 cagatgcgaa gttaagtgcg cagaaagtaa tatcatgcgt caatcgtatg tgaatgctgg
     3541 tcgctatact gctgtcgatt cgatactaac gccgccatcc agtgtcgaaa acgagctcga
     3601 attcatcgat gatatcagat ccactagtgg cctatgcggc cgctctagaa ctagtggatc
     3661 ccccgggctg caggaattcg atatcaagct tatcgatacc gtcgacctcg agggggggcc
     3721 cggtacccag cttttgttcc ctttagtgag ggttaattcc gagcttggcg taatcatggt
     3781 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac ataggagccg
     3841 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag gtaactcaca ttaattgcgt
     3901 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
     3961 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
     4021 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     4081 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     4141 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctcggccccc
     4201 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     4261 aaagatacca ggcgttcccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     4321 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcaatgct
     4381 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     4441 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     4501 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     4561 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     4621 ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     4681 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     4741 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     4801 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     4861 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     4921 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     4981 gtctatttcg ttcatccata gttgcctgac tgcccgtcgt gtagataact acgatacggg
     5041 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     5101 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     5161 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     5221 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     5281 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     5341 ccatgttgtg aaaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     5401 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     5461 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     5521 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     5581 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     5641 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     5701 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     5761 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     5821 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     5881 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gggtcctttt
     5941 catcacgtgc tataaaaata attataattt aaatttttta atataaatat ataaattaaa
     6001 aatagaaagt aaaaaaagaa attaaagaaa aaatagtttt tgttttccga agatgtaaaa
     6061 gactctaggg ggatcgccaa caaatactac cttttatctt gctcttcctg ctctcaggta
     6121 ttaatgccga attgtttcat cttgtctgtg tagaagacca cacacgaaaa tcctgtgatt
     6181 ttacatttta cttatcgtta atcgaatgta tatctattta atctgctttt cttgtctaat
     6241 aaatatatat gtaaagtacg ctttttgttg aaatttttta aacctttgtt tatttttttt
     6301 tcttcattcc gtaactcttc taccttcttt atttactttc taaaatccaa atacaaaaca
     6361 taaaaataaa taaacacaga gtaaattccc aaattattcc atcattaaaa gatacgaggc
     6421 gcgtgtaagt tacaggcaag cgatccgtcc taagaaacca ttattatcat gacattaacc
     6481 tataaaaata ggcgtatcac gaggcccttt cgtc
//
