LOCUS       sequence-56697-e        6049 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 6049)
  AUTHORS   Hughes AL, Gottschling DE
  TITLE     An early age increase in vacuolar pH limits mitochondrial function 
            and lifespan in yeast.
  JOURNAL   Nature. 2012 Dec 13;492(7428):261-5. doi: 10.1038/nature11654. Epub 
            2012 Nov 21.
  PUBMED    23172144
REFERENCE   2  (bases 1 to 6049)
  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: 
            "10.1038/nature11654"; journalName: "Nature"; date: "2012-12-13- 
            13"; volume: "492"; issue: "7428"; pages: "261-5"
COMMENT     Sequence Label: pAG306-GPD-empty chr I
FEATURES             Location/Qualifiers
     source          1..6049
                     /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        190..410
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             411..1214
                     /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(1343..1798)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1480..1501)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1692..1711
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1928..1950
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1942..1959
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1943..1959
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1966..1985
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1966..1984
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        2009..2653
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     2547..2570
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     primer_bind     complement(2676..2692)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2677..2693)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     terminator      2695..2942
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(2711..2729)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     promoter        complement(2979..2997)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3018..3034)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3018..3034)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3031..3053)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3042..3058
                     /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(3066..3072,3073..3090,3091..3096))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3111..3132
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3249..3266)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             complement(join(4079..4870,4871..4939))
                     /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     4702..4721
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4940..5044)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6022..6040
                     /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 accacgcttt tcaattcaat tcatcatttt ttttttattc ttttttttga tttcggtttc
      241 tttgaaattt ttttgattcg gtaatctccg aacagaagga agaacgaagg aaggagcaca
      301 gacttagatt ggtatatata cgcatatgta gtgttgaaga aacatgaaat tgcccagtat
      361 tcttaaccca actgcacaga acaaaaacct gcaggaaacg aagataaatc atgtcgaaag
      421 ctacatataa ggaacgtgct gctactcatc ctagtcctgt tgctgccaag ctatttaata
      481 tcatgcacga aaagcaaaca aacttgtgtg cttcattgga tgttcgtacc accaaggaat
      541 tactggagtt agttgaagca ttaggtccca aaatttgttt actaaaaaca catgtggata
      601 tcttgactga tttttccatg gagggcacag ttaagccgct aaaggcatta tccgccaagt
      661 acaatttttt actcttcgaa gacagaaaat ttgctgacat tggtaataca gtcaaattgc
      721 agtactctgc gggtgtatac agaatagcag aatgggcaga cattacgaat gcacacggtg
      781 tggtgggccc aggtattgtt agcggtttga agcaggcggc agaagaagta acaaaggaac
      841 ctagaggcct tttgatgtta gcagaattgt catgcaaggg ctccctatct actggagaat
      901 atactaaggg tactgttgac attgcgaaga gcgacaaaga ttttgttatc ggctttattg
      961 ctcaaagaga catgggtgga agagatgaag gttacgattg gttgattatg acacccggtg
     1021 tgggtttaga tgacaaggga gacgcattgg gtcaacagta tagaaccgtg gatgatgtgg
     1081 tctctacagg atctgacatt attattgttg gaagaggact atttgcaaag ggaagggatg
     1141 ctaaggtaga gggtgaacgt tacagaaaag caggctggga agcatatttg agaagatgcg
     1201 gccagcaaaa ctaaaaaact gtattataag taaatgcatg tatactaaac tcacaaatta
     1261 gagcttcaat ttaattatat cagttattac cctgcggtgt gaaataccgc acagatgcgt
     1321 aaggagaaaa taccgcatca ggaaattgta aacgttaata ttttgttaaa attcgcgtta
     1381 aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat
     1441 aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca
     1501 ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc
     1561 ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta
     1621 aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg
     1681 gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg
     1741 gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcg
     1801 cgccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg
     1861 ctattacgcc agctggcgaa ggggggatgt gctgcaaggc gattaagttg ggtaacgcca
     1921 gggttttccc agtcacgacg ttgtaaaacg acggccagtg aattgtaata cgactcacta
     1981 tagggcgaat tggagctctt cgagtttatc attatcaata ctcgccattt caaagaatac
     2041 gtaaataatt aatagtagtg attttcctaa ctttatttag tcaaaaaatt agccttttaa
     2101 ttctgctgta acccgtacat gcccaaaata gggggcgggt tacacagaat atataacacc
     2161 gaaggtgcct gggtgaacag gtttattcct ggcatccact aaatataatg gagcccgctt
     2221 tttaagctgg catccagaaa aaaaaagaat cccagcacca aaatattgtt ttcttcacca
     2281 accatcagtt cataggtcca ttctcttagc gcaactacag agaacagggg cacaaacagg
     2341 caaaaaacgg gcacaacctc aatggagtga tgcaacctgc ctggagtaaa tgatgacaca
     2401 aggcaattga cccacgcatg tatctatctc attttcttac accttctatt accttctgct
     2461 ctctctgatt tggaaaaagc tgaaaaaaaa ggttgaaacc agttccctga aattattccc
     2521 ctacttgact aataagtata taaagacggt aggtattgat tgtaattctg taaatctatt
     2581 tcttaaactt cttaaattct acttttatag ttagtctttt ttttagtttt aaaacaccag
     2641 aacttagttt cgacggattc tagaactagg cttatcgata ccgtcgacct cgagtcatgt
     2701 aattagttat gtcacgctta cattcacgcc ctccccccac atccgctcta accgaaaagg
     2761 aaggagttag acaacctgaa gtctaggtcc ctatttattt ttttatagtt atgttagtat
     2821 taagaacgtt atttatattt caaatttttc ttttttttct gtacagacgc gtgtacgcat
     2881 gtaacattat actgaaaacc ttgcttgaga aggttttggg acgctcgaag gctttaattt
     2941 gcggccgtcg agggggggcc cggtaccagc ttttgttccc tttagtgagg gttaatttcg
     3001 agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
     3061 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
     3121 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     3181 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
     3241 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     3301 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     3361 atgtgagcaa aaggccagca aaaggccagg caggactata aagataccag gcgttccccc
     3421 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     3481 cctttctccc ttcgggaagc gtggcgcttt ctcaatgctc acgctgtagg tatctcagtt
     3541 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     3601 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     3661 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     3721 agttcttgaa gtggtggcct aactacggct acactagaag gacagtattt ggtatctgcg
     3781 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     3841 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     3901 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     3961 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     4021 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     4081 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     4141 ttgcctgact gcccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     4201 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     4261 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     4321 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     4381 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     4441 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtga aaaaaagcgg
     4501 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     4561 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     4621 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     4681 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     4741 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     4801 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     4861 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     4921 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     4981 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     5041 cgcgcacatt tccccgaaaa gtgccacctg gggggtgcta attatggcat tgattggggc
     5101 ggtaaaacat tcaaagtcaa tgaaaaggat gctaaattga atgatactga agacatttca
     5161 ttcgcttctc agatcaaaaa cttaaatctg tctatatgag ctgtcccaaa gccacttctt
     5221 acctttcaag aaaacatgga ccttgctttg ccacaaaata attgcctgta agatagtttt
     5281 ttatttgcac attgttatta ctaacctatg ctttgctacg gacgactgct tggtaaaaat
     5341 gcaaaagtat taggatgtct cacaagaaac tgctcgtggt gaaaattttc agccgcgacc
     5401 atcctcccac gccaacaagc cctcttttgc ccatccttct ggaattcgcg gtgaaaatca
     5461 atttgatata taaaattccc tcataatatc taggaacggg atcacatatc tcaatgagct
     5521 aaagcggccg cttcttcgaa tgggaccagc taaagtgatc aaacttcatg caggatgttg
     5581 gacacttttt ctttcaagaa gctccaaata tataaatttt tgcaatatag gaattgaaat
     5641 atcgtacttc cgattcagaa tatctcaact ctactacatc tatatatgac tgtactcgcg
     5701 ttcttggtaa acatcaatta gtaggtgttt ttgcgagcat atgttgagct cagaaaatat
     5761 gagcacaatc ttcgaaacca ttgttttttt gtgcaaaagt caccatcggg tttatagtat
     5821 ggactcttga acgaaccatt ctgatctgta tttctctcat atttactcca tttatttttt
     5881 gctgctcttt gctttacttc aacacgatgc ttttcctaag taatttttta ttttattcac
     5941 aatgctccat aactcttcag gccttaaacg accatccatg gcccctaaga aaccattatt
     6001 atcatgacat taacctataa aaataggcgt atcacgaggc cctttcgtc
//
