LOCUS       sequence-120917-        5186 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  URA3 vector for cloning guide sequences using BclI-SwaI sites into 
            guide RNA expression cassette.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5186)
  AUTHORS   Laughery MF, Hunter T, Brown A, Hoopes J, Ostbye T, Shumaker T, 
            Wyrick JJ
  TITLE     New vectors for simple and streamlined CRISPR-Cas9 genome editing in
            Saccharomyces cerevisiae.
  JOURNAL   Yeast. 2015 Dec;32(12):711-20. doi: 10.1002/yea.3098. Epub 2015 Sep 
            21.
  PUBMED    26305040
REFERENCE   2  (bases 1 to 5186)
  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"; doi: "10.1002/yea.3098"; 
            journalName: "Yeast"; date: "2015-12"; volume: "32"; issue: "12"; 
            pages: "711-20"
COMMENT     Sequence Label: pT040
FEATURES             Location/Qualifiers
     source          1..5186
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        27..131
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(132..200,201..992)
                     /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(350..369)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1163..1751
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1652..1671
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1905..1922
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2039..2060
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2075..2080,2081..2098,2099..2105)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2113..2129
                     /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     2118..2140
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2137..2153
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2137..2153
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        2174..2192
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     2224..2240
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     2225..2241
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2287..2303)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2287..2303)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     terminator      complement(2378..2397)
                     /label=SUP4 terminator
                     /note="transcription terminator for the S. cerevisiae SUP4 
                     tRNA gene"
     promoter        complement(2479..2747)
                     /label=SNR52 promoter
                     /note="promoter for the S. cerevisiae small nucleolar RNA 
                     gene SNR52"
     primer_bind     complement(2773..2792)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2774..2792)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2799..2816)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2799..2815)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2808..2830)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     misc_feature    3331..3623
                     /label=CEN4
                     /note="S. cerevisiae CEN4 centromere"
     rep_origin      3624..4077
                     /label=ARS1
                     /note="S. cerevisiae autonomously replicating sequence 
                     ARS1"
     promoter        4084..4304
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             4305..5108
                     /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"
ORIGIN
        1 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
       61 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
      121 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
      181 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
      241 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
      301 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
      361 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
      421 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
      481 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
      541 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
      601 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
      661 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
      721 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
      781 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
      841 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
      901 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
      961 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     1021 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     1081 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     1141 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     1201 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     1261 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt
     1321 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     1381 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     1441 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     1501 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     1561 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     1621 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     1681 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     1741 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     1801 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     1861 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
     1921 aggaagcgga agagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt
     1981 aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta
     2041 atgtgagtta gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta
     2101 tgttgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagctat gaccatgatt
     2161 acgccaagct cgaaattaac cctcactaaa gggaacaaaa gctggtaccg ggccccccct
     2221 cgatcgaggt cgacggtatc gataagcttg atatcgaatt cctgcagccc ccgctcgagc
     2281 gggggggatc cactagttct agagcggccg tggtatcgtt tagattggca attacagtgt
     2341 cttagctcac atgcttataa ctaattacat gactcgaaga cataaaaaac aaaaaaagca
     2401 ccaccgactc ggtgccactt tttcaagttg ataacggact agccttattt taacttgcta
     2461 tttaaatgca tcgattctga tcatttatct ttcactgcgg agaagtttcg aacgccgaaa
     2521 catgcgcacc aactttcact tctacagcgt ttgaccaaaa tcttttgaac agaacattgt
     2581 agggtgtgaa aaaatgcgca cctttaccgc tagcccaaga gggcactaca aaatctagag
     2641 ttgtacttca aacgtacatg taatcacctt gtatatactc gaaagaaaac atcaagtttc
     2701 tgtataaata tgagtgaaag cataatcata cattatcttt tcaaagattg tccgcggtgg
     2761 agctccaatt cgccctatag tgagtcgtat tacaattcac tggccgtcgt tttacaacgt
     2821 cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc
     2881 gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc
     2941 ctgaatggcg aatggcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca
     3001 caccgcataa ctagctttta acttgtatcc taggttatct atgctgtctc accatagaga
     3061 atattaccta tttcagaatg tatgtccatg attcgccggg taaatacata taatacacaa
     3121 atctggctta ataaagtcta taatatatct cataaagaag tgctaaattg gctagtgcta
     3181 tatattttta agaaaatttc ttttgactaa gtccatatcg actttgtaaa agttcacttt
     3241 agcatacata tattacacga gccagaaatt gtaacttttg cctaaaatca caaattgcaa
     3301 aatttaattg cttgcaaaag gtcacatgct tataatcaac ttttttaaaa atttaaaata
     3361 cttttttatt ttttattttt aaacataaat gaaataattt atttattgtt tatgattacc
     3421 gaaacataaa acctgctcaa gaaaaagaaa ctgttttgtc cttggaaaaa aagcactacc
     3481 taggagcggc caaaatgccg aggctttcat agcttaaact ctttacagaa aataggcatt
     3541 atagatcagt tcgagttttc ttattcttcc ttccggtttt atcgtcacag ttttacagta
     3601 aataagtatc acctcttaga gttggcggtt gtttgcaaga ccgagaaaag gctagcaaga
     3661 atcgggtcat tgtagcgtat gcgcctgtga acattctctt caacaagttt gattccattg
     3721 cggtgaaatg gtaaaagtca accccctgcg atgtatattt tcctgtacaa tcaatcaaaa
     3781 agccaaatga tttagcatta tctttacatc ttgttatttt acagatttta tgtttagatc
     3841 ttttatgctt gcttttcaaa aggcctgcag gcaagtgcac aaacaatact taaataaata
     3901 ctactcagta ataacctatt tcttagcatt tttgacgaaa tttgctattt tgttagagtc
     3961 ttttacacca tttgtctcca cacctccgct tacatcaaca ccaataacgc catttaatct
     4021 aagcgcatca ccaacatttt ctggcgtcag tccaccagct aacataaaat gtaagctaag
     4081 cttttcaatt caattcatca tttttttttt attctttttt ttgatttcgg tttctttgaa
     4141 atttttttga ttcggtaatc tccgaacaga aggaagaacg aaggaaggag cacagactta
     4201 gattggtata tatacgcata tgtagtgttg aagaaacatg aaattgccca gtattcttaa
     4261 cccaactgca cagaacaaaa acctgcagga aacgaagata aatcatgtcg aaagctacat
     4321 ataaggaacg tgctgctact catcctagtc ctgttgctgc caagctattt aatatcatgc
     4381 acgaaaagca aacaaacttg tgtgcttcat tggatgttcg taccaccaag gaattactgg
     4441 agttagttga agcattaggt cccaaaattt gtttactaaa aacacatgtg gatatcttga
     4501 ctgatttttc catggagggc acagttaagc cgctaaaggc attatccgcc aagtacaatt
     4561 ttttactctt cgaagacaga aaatttgctg acattggtaa tacagtcaaa ttgcagtact
     4621 ctgcgggtgt atacagaata gcagaatggg cagacattac gaatgcacac ggtgtggtgg
     4681 gcccaggtat tgttagcggt ttgaagcagg cggcagaaga agtaacaaag gaacctagag
     4741 gccttttgat gttagcagaa ttgtcatgca agggctccct atctactgga gaatatacta
     4801 agggtactgt tgacattgcg aagagcgaca aagattttgt tatcggcttt attgctcaaa
     4861 gagacatggg tggaagagat gaaggttacg attggttgat tatgacaccc ggtgtgggtt
     4921 tagatgacaa gggagacgca ttgggtcaac agtatagaac cgtggatgat gtggtctcta
     4981 caggatctga cattattatt gttggaagag gactatttgc aaagggaagg gatgctaagg
     5041 tagagggtga acgttacaga aaagcaggct gggaagcata tttgagaaga tgcggccagc
     5101 aaaactaaaa aactgtatta taagtaaatg catgtatact aaactcacaa attagagctt
     5161 caatttaatt atatcagtta ttaccc
//
