LOCUS       sequence-104893-        5887 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pRS416 backbone, contains an RFP flanked by BsaI sites with CAGT.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5887)
  AUTHORS   Agmon N, Mitchell LA, Cai Y, Ikushima S, Chuang J, Zheng A, Choi WJ,
            Martin JA, Caravelli K, Stracquadanio G, Boeke JD
  TITLE     Yeast Golden Gate (yGG) for the Efficient Assembly of S. cerevisiae 
            Transcription Units.
  JOURNAL   ACS Synth Biol. 2015 Jul 17;4(7):853-9. doi: 10.1021/sb500372z. Epub
            2015 Mar 23.
  PUBMED    25756291
REFERENCE   2  (bases 1 to 5887)
  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: "10.1021/sb500372z";
            journalName: "ACS Synth Biol"; date: "2015-07-17- 17"; volume: "4"; 
            issue: "7"; pages: "853-9"
COMMENT     Sequence Label: pAV116.loxP
FEATURES             Location/Qualifiers
     source          1..5887
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     27..45
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     145..164
                     /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(1345..1800)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1482..1503)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1694..1713
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1930..1952
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1944..1961
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1945..1961
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1971..1990
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1971..1989
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    2035..2068
                     /label=loxPsym
                     /bound_moiety="Cre recombinase"
                     /note="loxPsym has a symmetric spacer region, and loxPsym 
                     sites are capable of recombining in either orientation 
                     (Hoess et al., 1986)."
     CDS             complement(2148..2825)
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     RBS             2832..2843
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     protein_bind    2861..2877
                     /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(2885..2891,2892..2909,2910..2915))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2930..2951
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     protein_bind    3081..3114
                     /label=loxPsym
                     /bound_moiety="Cre recombinase"
                     /note="loxPsym has a symmetric spacer region, and loxPsym 
                     sites are capable of recombining in either orientation 
                     (Hoess et al., 1986)."
     primer_bind     complement(3167..3187)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3167..3185)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3206..3222)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3206..3222)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     protein_bind    3230..3246
                     /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(3254..3260,3261..3278,3279..3284))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(3437..3454)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3608..4196)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3688..3707)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4367..5158,5159..5227))
                     /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     4990..5009
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5228..5332)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    5369..5872
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
ORIGIN
        1 ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt gtcacagctt
       61 gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg
      121 ggtgtcgggg ctggcttaac tatgcggcat cagagcagat tgtactgaga gtgcaccata
      181 ccacagcttt 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 gatgcattgg gtcaacagta tagaaccgtg gatgatgtgg
     1081 tgtctacagg atctgacatt attattgttg gaagaggact atttgcaaag ggaagggatg
     1141 ctaaggtaga gggtgaacgt tacagaaaag caggctggga agcatatttg agaagatgcg
     1201 gccagcaaaa ctaaaaaact gtattataag taaatgcatg tatactaaac tcacaaatta
     1261 gagcttcaat ttaattatat cagttattac cctatgcggt gtgaaatacc gcacagatgc
     1321 gtaaggagaa aataccgcat caggaaattg taaacgttaa tattttgtta aaattcgcgt
     1381 taaatttttg ttaaatcagc tcatttttta accaataggc cgaaatcggc aaaatccctt
     1441 ataaatcaaa agaatagacc gagatagggt tgagtgttgt tccagtttgg aacaagagtc
     1501 cactattaaa gaacgtggac tccaacgtca aagggcgaaa aaccgtctat cagggcgatg
     1561 gcccactacg tgaaccatca ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac
     1621 taaatcggaa ccctaaaggg agcccccgat ttagagcttg acggggaaag ccggcgaacg
     1681 tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag
     1741 cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt
     1801 cgcgccattc gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt
     1861 cgctattacg ccagctggcg aaagggggat gtgctgcaag gcgattaagt tgggtaacgc
     1921 cagggttttc ccagtcacga cgttgtaaaa cgacggccag tgagcgcgcg taatacgact
     1981 cactataggg cgaattgggt accgggcccc ccctcgaggt cgacggcagc tgggataact
     2041 tcgtataatg tacattatac gaagttatgg gacgtggcga attcctgcag cccccagtag
     2101 agaccgcctg gctctagtag cgatctacac tagcactatc agcgttatta agcaccggtg
     2161 gagtgacgac cttcagcacg ttcgtactgt tcaacgatgg tgtagtcttc gttgtgggag
     2221 gtgatgtcca gtttgatgtc ggttttgtaa gcacccggca gctgaaccgg ttttttagcc
     2281 atgtaggtgg ttttaacttc agcgtcgtag tgaccaccgt ctttcagttt cagacgcatt
     2341 ttgatttcac ctttcagagc accgtcttcc gggtacatac gttcggtgga agcttcccaa
     2401 cccatggttt ttttctgcat aaccggaccg tcggacggga agttggtacc acgcagttta
     2461 actttgtaga tgaactcacc gtcttgcagg gaggagtcct gggtaacggt aacaacacca
     2521 ccgtcttcga agttcataac acgttcccat ttgaaacctt ccgggaagga cagtttcagg
     2581 tagtccggga tgtcagccgg gtgtttaacg taagctttgg aaccgtactg gaactgcggg
     2641 gacaggatgt cccaagcgaa cggcagcgga ccacctttgg taactttcag tttagcggtc
     2701 tgggtacctt cgtacggacg accttcacct tcaccttcga tttcgaactc gtgaccgtta
     2761 acggaacctt ccatacgaac tttgaaacgc atgaactctt tgataacgtc ttcggaggaa
     2821 gccatctagt atttctcctc tttctctagt atgtgtgaaa ttgttatccg ctcacaattc
     2881 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct
     2941 aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
     3001 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggtctcatt
     3061 ttggggatcc cggagcaccc ataacttcgt ataatgtaca ttatacgaag ttatcccagg
     3121 tcgctctaga gcggccgcca ccgcggtgga gctccagctt ttgttccctt tagtgagggt
     3181 taattgcgcg cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
     3241 tcacaattcc acacaacata ggagccggaa gcataaagtg taaagcctgg ggtgcctaat
     3301 gagtgaggta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     3361 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     3421 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     3481 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     3541 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     3601 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     3661 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     3721 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     3781 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     3841 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     3901 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     3961 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     4021 ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc
     4081 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     4141 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     4201 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     4261 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     4321 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     4381 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     4441 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     4501 taccgcgaga tccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     4561 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     4621 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     4681 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     4741 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     4801 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     4861 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     4921 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     4981 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     5041 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     5101 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     5161 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     5221 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     5281 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     5341 cccgaaaagt gccacctggg tccttttcat cacgtgctat aaaaataatt ataatttaaa
     5401 ttttttaata taaatatata aattaaaaat agaaagtaaa aaaagaaatt aaagaaaaaa
     5461 tagtttttgt tttccgaaga tgtaaaagac tctaggggga tcgccaacaa atactacctt
     5521 ttatcttgct cttcctgctc tcaggtatta atgccgaatt gtttcatctt gtctgtgtag
     5581 aagaccacac acgaaaatcc tgtgatttta cattttactt atcgttaatc gaatgtatat
     5641 ctatttaatc tgcttttctt gtctaataaa tatatatgta aagtacgctt tttgttgaaa
     5701 ttttttaaac ctttgtttat ttttttttct tcattccgta actcttctac cttctttatt
     5761 tactttctaa aatccaaata caaaacataa aaataaataa acacagagta aattcccaaa
     5821 ttattccatc attaaaagat acgaggcgcg tgtaagttac aggcaagcga tccgtcctaa
     5881 gaaacca
//
