LOCUS       sequence-350313-        4986 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Yeast shuttle plasmid that allows conversion between low copy, high 
            copy, and integrating vectors. Low copy, URA3-selected..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4986)
  AUTHORS   Nickerson DP, Quinn MA, Milnes JM
  TITLE     Rapid conversion of replicating and integrating Saccharomyces 
            cerevisiae plasmid vectors via Cre recombinase.
  JOURNAL   G3 (Bethesda). 2021 Dec 8;11(12). pii: 6380430. doi: 
            10.1093/g3journal/jkab336.
  PUBMED    34599813
REFERENCE   2  (bases 1 to 4986)
  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: "G3 
            (Bethesda)."; date: "2021-12-8"; volume: "11(12). pii: 6380430. 
            doi"; pages: " 10.1093/g3journal/jkab336"
COMMENT     Sequence Label: pDN616
FEATURES             Location/Qualifiers
     source          1..4986
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    468..501
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     539..557
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(595..617)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     717..736
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        760..975
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             976..1779
                     /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"
     rep_origin      complement(1910..2365)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2047..2068)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2259..2278
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     CDS             complement(2342..2722)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWSSTAVAAALELVDPPGCRNSISSLSI
                     PSTSRGGPVPNSPYSESYYARSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEAR
                     TDRPSQQLRSLNGEWTRPVAAH"
     primer_bind     2491..2513
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2505..2522
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2506..2522
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2532..2551
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2532..2550
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    2559..2666
                     /label=MCS
                     /note="pBluescript multiple cloning site"
     primer_bind     2575..2591
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     2576..2592
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2626..2642)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2626..2642)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(2679..2699)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2679..2697)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2718..2734)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2718..2734)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2731..2753)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2742..2758
                     /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(2766..2772,2773..2790,2791..2796))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2811..2832
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2949..2966)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3120..3708)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3200..3219)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3879..4670,4671..4739))
                     /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     4502..4521
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4740..4844)
                     /gene="bla"
                     /label=AmpR promoter
     protein_bind    4870..4903
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    join(4939..4986,1..456)
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
ORIGIN
        1 tataaattaa aaatagaaag taaaaaaaga aattaaagaa aaaatagttt ttgttttccg
       61 aagatgtaaa agactctagg gggatcgcca acaaatacta ccttttatct tgctcttcct
      121 gctctcaggt attaatgccg aattgtttca tcttgtctgt gtagaagacc acacacgaaa
      181 atcctgtgat tttacatttt acttatcgtt aatcgaatgt atatctattt aatctgcttt
      241 tcttgtctaa taaatatata tgtaaagtac gctttttgtt gaaatttttt aaacctttgt
      301 ttattttttt ttcttcattc cgtaactctt ctaccttctt tatttacttt ctaaaatcca
      361 aatacaaaac ataaaaataa ataaacacag agtaaattcc caaattattc catcattaaa
      421 agatacgagg cgcgtgtaag ttacaggcaa gcgatccgtc cgacgtcata acttcgtata
      481 atgtatgcta tacgaagtta tttaagaaac cattattatc atgacattaa cctataaaaa
      541 taggcgtatc acgaggccct ttcgtctcgc gcgtttcggt gatgacggtg aaaacctctg
      601 acacatgcag ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca
      661 agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg ggctggctta actatgcggc
      721 atcagagcag attgtactga gagtgcacca taccaccttt tcaattcatc attttttttt
      781 tattcttttt tttgatttcg gtttccttga aatttttttg attcggtaat ctccgaacag
      841 aaggaagaac gaaggaagga gcacagactt agattggtat atatacgcat atgtagtgtt
      901 gaagaaacat gaaattgccc agtattctta acccaactgc acagaacaaa aacctgcagg
      961 aaacgaagat aaatcatgtc gaaagctaca tataaggaac gtgctgctac tcatcctagt
     1021 cctgttgctg ccaagctatt taatatcatg cacgaaaagc aaacaaactt gtgtgcttca
     1081 ttggatgttc gtaccaccaa ggaattactg gagttagttg aagcattagg tcccaaaatt
     1141 tgtttactaa aaacacatgt ggatatcttg actgattttt ccatggaggg cacagttaag
     1201 ccgctaaagg cattatccgc caagtacaat tttttactct tcgaagacag aaaatttgct
     1261 gacattggta atacagtcaa attgcagtac tctgcgggtg tatacagaat agcagaatgg
     1321 gcagacatta cgaatgcaca cggtgtggtg ggcccaggta ttgttagcgg tttgaagcag
     1381 gcggcagaag aagtaacaaa ggaacctaga ggccttttga tgttagcaga attgtcatgc
     1441 aagggctccc tatctactgg agaatatact aagggtactg ttgacattgc gaagagcgac
     1501 aaagattttg ttatcggctt tattgctcaa agagacatgg gtggaagaga tgaaggttac
     1561 gattggttga ttatgacacc cggtgtgggt ttagatgaca agggagacgc attgggtcaa
     1621 cagtatagaa ccgtggatga tgtggtctct acaggatctg acattattat tgttggaaga
     1681 ggactatttg caaagggaag ggatgctaag gtagagggtg aacgttacag aaaagcaggc
     1741 tgggaagcat atttgagaag atgcggccag caaaactaaa aaactgtatt ataagtaaat
     1801 gcatgtatac taaactcaca aattagagct tcaatttaat tatatcagtt attaccctat
     1861 gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac cgcatcagga aattgtaagc
     1921 gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa
     1981 taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt
     2041 gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg
     2101 cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt
     2161 ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga
     2221 gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg
     2281 ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg
     2341 cttaatgcgc cgctacaggg cgcgtccatt cgccattcag gctgcgcaac tgttgggaag
     2401 ggcgatcggt gcgggcctct tcgctattac gccagctggc gaaaggggga tgtgctgcaa
     2461 ggcgattaag ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca
     2521 gtgagcgcgc gtaatacgac tcactatagg gcgaattggg taccgggccc cccctcgagg
     2581 tcgacggtat cgataagctt gatatcgaat tcctgcagcc cgggggatcc actagttcta
     2641 gagcggccgc caccgcggtg gagctccagc ttttgttccc tttagtgagg gttaattgcg
     2701 cgcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
     2761 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
     2821 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     2881 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
     2941 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     3001 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     3061 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     3121 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     3181 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     3241 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     3301 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     3361 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     3421 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     3481 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     3541 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc
     3601 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     3661 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     3721 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     3781 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
     3841 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
     3901 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
     3961 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
     4021 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
     4081 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
     4141 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     4201 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     4261 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     4321 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     4381 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     4441 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
     4501 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
     4561 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
     4621 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
     4681 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
     4741 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac
     4801 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa
     4861 gtgccaccta taacttcgta taatgtatgc tatacgaagt tattgacgtc cccgaaaagt
     4921 gccacctggg tccttttcat cacgtgctat aaaaataatt ataatttaaa ttttttaata
     4981 taaata
//
