LOCUS       sequence-104906-        6353 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pCACS backbone (Amp), LEU2, 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 6353)
  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 6353)
  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: pAV10.HO5.loxP
FEATURES             Location/Qualifiers
     source          1..6353
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(61..78)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(232..820)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(312..331)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(1003..1097)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             join(1115..1183,1184..1975)
                     /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(1333..1352)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     primer_bind     2146..2164
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(2168..2211)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     promoter        2960..3367
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     CDS             3368..4462
                     /codon_start=1
                     /gene="S. cerevisiae LEU2"
                     /product="3-isopropylmalate dehydrogenase, required for 
                     leucine biosynthesis"
                     /label=LEU2
                     /note="yeast auxotrophic marker"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKADAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVNPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     protein_bind    4762..4795
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(4849..5526)
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     RBS             5533..5544
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     protein_bind    5562..5578
                     /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(5586..5592,5593..5610,5611..5616))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5631..5652
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     protein_bind    5764..5797
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     terminator      join(6333..6353,1..37)
                     /label=his operon terminator
                     /note="This putative transcriptin terminator from the E. 
                     coli his operon has a 2-bp deletion introduced during 
                     synthesis. Its efficiency has not been determined."
ORIGIN
        1 gtcaccaccc tgcccttttt ctttaaaacc gaaaagatta cttcgcgtta tgcaggcttc
       61 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
      121 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
      181 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccacag
      241 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
      301 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
      361 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
      421 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
      481 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
      541 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
      601 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
      661 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
      721 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
      781 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
      841 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
      901 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
      961 aagtatatat gagtaaactt ggtctgacag ctcgaggctt ggattctcac caataaaaaa
     1021 cgcccggcgg caaccgagcg ttctgaacaa atccagatgg agttctgagg tcattactgg
     1081 atctatcaac aggagtccaa gcgagctcga tatcatgagt attcaacatt tccgtgtcgc
     1141 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt
     1201 gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct
     1261 caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac
     1321 ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact
     1381 cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa
     1441 gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga
     1501 taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt
     1561 tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga
     1621 agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg
     1681 caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat
     1741 ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat
     1801 tgctgataaa tctggagccg gtgagcgtgg atctcgcggt atcattgcag cactggggcc
     1861 agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga
     1921 tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaatttct
     1981 ccattttagc ttccttagct cctgaaaatc tcgataactc aaaaaatacg cccggtagtg
     2041 atcttatttc attatggtga aagttggaac ctcttacgtg cccgatcaac tcgagtgcca
     2101 cctgacgtct aagaaaccat tattatcatg acattaacct ataaaaatag gcgtatcacg
     2161 aggcagaatt tcagataaaa aaaatcctta gctttcgcta aggatgattt ctggaattcg
     2221 cggccgcgta tccttgccta acttcaatgt caaagttgaa atattctcct ttagagcgct
     2281 ccatttcttc tatgaagcgt tttgcggcaa actcaccttc aactgtcatt gggaatgtct
     2341 tatgatggtt ttttggaatt attattatcc taccatcaag cgtctgacat tgctgcagat
     2401 ttctccatct cactttatat ttggtggcat ttctaccact tttttccaac agtggtttgg
     2461 tagggaccct gactgacaat ttatgacctg cagtacattg taatgcaaga cgctgataaa
     2521 ctgttctacg cctgggatct aacctaccag gttcaccttc aaaagctctg tgtttggttt
     2581 tttgctgtat attatagatt ttctgatagc cctgtgtgac atttatgacg cgggcagcgg
     2641 agccatctgc gcacataacg taagagttag ccgtgacgtt tgcgatgtct ttaatttcac
     2701 cgttagccat cagaatagtc gtgttttcag aaagcatgga tccacatacc taatattatt
     2761 gccttattaa aaatggaatc ccaacaatta catcaaaatc cacattctct tcaaaatcaa
     2821 ttgtcctgta cttccttgtt catgtgtgtt caaaaacgtt atatttatag gataattata
     2881 ctctatttct caacaagtaa ttggttgttt ggccgagcgg tctaaggcgc ctgattcaag
     2941 aaatatcttg accgcagtta actgtgggaa tactcaggta tcgtaagatg caagagttcg
     3001 aatctcttag caaccattat ttttttcctc aacataacga gaacacacag gggcgctatc
     3061 gcacagaatc aaattcgatg actggaaatt ttttgttaat ttcagaggtc gcctgacgca
     3121 tatacctttt tcaactgaaa aattgggaga aaaaggaaag gtgagagcgc cggaaccggc
     3181 ttttcatata gaatagagaa gcgttcatga ctaaatgctt gcatcacaat acttgaagtt
     3241 gacaatatta tttaaggacc tattgttttt tccaataggt ggttagcaat cgtcttactt
     3301 tctaactttt cttacctttt acatttcagc aatatatata tatatatttc aaggatatac
     3361 cattctaatg tctgccccta agaagatcgt cgttttgcca ggtgaccacg ttggtcaaga
     3421 aatcacagcc gaagccatta aggttcttaa agctatttct gatgttcgtt ccaatgtcaa
     3481 gttcgatttc gaaaatcatt taattggtgg tgctgctatc gatgctacag gtgttccact
     3541 tccagatgag gcgctggaag cctccaagaa ggctgatgcc gttttgttag gtgctgtggg
     3601 tggtcctaaa tggggtaccg gtagtgttag acctgaacaa ggtttactaa aaatccgtaa
     3661 agaacttcaa ttgtacgcca acttaagacc atgtaacttt gcatccgact ctcttttaga
     3721 cttatctcca atcaagccac aatttgctaa aggtactgac ttcgttgttg tcagagaatt
     3781 agtgggaggt atttactttg gtaagagaaa ggaagacgat ggtgatggtg tcgcttggga
     3841 tagtgaacaa tacaccgttc cagaagtgca aagaatcaca agaatggccg ctttcatggc
     3901 cctacaacat gagccaccat tgcctatttg gtccttggat aaagctaatg ttttggcctc
     3961 ttcaagatta tggagaaaaa ctgtggagga aaccatcaag aacgaattcc ctacattgaa
     4021 ggttcaacat caattgattg attctgccgc catgatccta gttaagaacc caacccacct
     4081 aaatggtatt ataatcacca gcaacatgtt tggtgatatc atctccgatg aagcctccgt
     4141 tatcccaggt tccttgggtt tgttgccatc tgcgtccttg gcctctttgc cagacaagaa
     4201 caccgcattt ggtttgtacg aaccatgcca cggttctgct ccagatttgc caaagaataa
     4261 ggtcaaccct atcgccacta tcttgtctgc tgcaatgatg ttgaaattgt cattgaactt
     4321 gcctgaagaa ggtaaggcca ttgaagatgc agttaaaaag gttttggatg caggtatcag
     4381 aactggtgat ttaggtggtt ccaacagtac caccgaagtc ggtgatgctg tcgccgaaga
     4441 agttaagaaa atccttgctt aaaaagattc tcttttttta tgatatttgt acataaactt
     4501 tataaatgaa attcataata gaaacgacac gaaattacaa aatggaatat gttcataggg
     4561 tagacgaaac tatatacgca atctacatac atttatcaag aaggagaaaa aggaggatgt
     4621 aaaggaatac aggtaagcaa attgatacta atggctcaac gtgataagga aaaagaattg
     4681 cactttaaca ttaatattga caaggaggag ggcaccacac aaaaagttag gtgtaacaga
     4741 aaatcatgaa actagctagc aataacttcg tataatgtat gctatacgaa gttatcagta
     4801 gagaccgcct ggctctagta gcgatctaca ctagcactat cagcgttatt aagcaccggt
     4861 ggagtgacga ccttcagcac gttcgtactg ttcaacgatg gtgtagtctt cgttgtggga
     4921 ggtgatgtcc agtttgatgt cggttttgta agcacccggc agctgaaccg gttttttagc
     4981 catgtaggtg gttttaactt cagcgtcgta gtgaccaccg tctttcagtt tcagacgcat
     5041 tttgatttca cctttcagag caccgtcttc cgggtacata cgttcggtgg aagcttccca
     5101 acccatggtt tttttctgca taaccggacc gtcggacggg aagttggtac cacgcagttt
     5161 aactttgtag atgaactcac cgtcttgcag ggaggagtcc tgggtaacgg taacaacacc
     5221 accgtcttcg aagttcataa cacgttccca tttgaaacct tccgggaagg acagtttcag
     5281 gtagtccggg atgtcagccg ggtgtttaac gtaagctttg gaaccgtact ggaactgcgg
     5341 ggacaggatg tcccaagcga acggcagcgg accacctttg gtaactttca gtttagcggt
     5401 ctgggtacct tcgtacggac gaccttcacc ttcaccttcg atttcgaact cgtgaccgtt
     5461 aacggaacct tccatacgaa ctttgaaacg catgaactct ttgataacgt cttcggagga
     5521 agccatctag tatttctcct ctttctctag tatgtgtgaa attgttatcc gctcacaatt
     5581 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
     5641 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     5701 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggtctcat
     5761 tttataactt cgtataatgt atgctatacg aagttatact agtgcatgct ttaaagtata
     5821 gatagaattg attgctgctt atgaggatat ggatttagag ctaatagttg taatagaaat
     5881 aaaagccatt tagaatagga attgaataca attttatgag ctttaaatat ttaatgatga
     5941 atagtgcgtt tagaacgctt catcatgctt caacaagctt ctgcaaacct catttgagta
     6001 gcaatagctg tttacttttt tccgcgactt tattcgaagt tatggtgaac ttttcgtggt
     6061 ttcacaccta ataacgccca gcggtagtac aattcaaagt agtaggtacc aatggtagta
     6121 ctagtgggtg atagtagcaa atttctctat gaaaatgaat tgttgctctg caatgatgtt
     6181 aatgtggaca tgacatgaat tttcgcgata tgttgatgta aataataaca tgattttcgt
     6241 ggatcatttc gtaaaccata ggtttatttc gtgaattact aagagaaagg catagaaaca
     6301 ggacttgcgg atgggatacg cggccgctgc agtccggcaa aaaagggcaa ggt
//
