LOCUS       sequence-104899-        5541 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pCACS backbone (Amp), SpHIS5, 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 5541)
  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 5541)
  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.K3.loxP
FEATURES             Location/Qualifiers
     source          1..5541
                     /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"
     gene            2708..3908
                     /label=HIS3MX6
                     /note="yeast selectable marker encoding the S. pombe his5 
                     gene, which corresponds to S. cerevisiae HIS3"
     promoter        2708..3051
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             3052..3705
                     /codon_start=1
                     /gene="Schizosaccharomyces pombe his5"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=S. pombe his5
                     /note="yeast auxotrophic marker; corresponds to S. 
                     cerevisiae HIS3"
                     /translation="MGRRAFVERNTNETKISVAIALDKAPLPEESNFIDELITSKHANQ
                     KGEQVIQVDTGIGFLDHMYHALAKHAGWSLRLYSRGDLIIDDHHTAEDTAIALGIAFKQ
                     AMGNFAGVKRFGHAYCPLDEALSRSVVDLSGRPYAVIDLGLKREKVGELSCEMIPHLLY
                     SFSVAAGITLHVTCLYGSNDHHRAESAFKSLAVAMRAATSLTGSSEVPSTKGVL"
     terminator      3711..3908
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     protein_bind    3947..3980
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(4034..4711)
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     RBS             4718..4729
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     protein_bind    4747..4763
                     /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(4771..4777,4778..4795,4796..4801))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4816..4837
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     protein_bind    4949..4982
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     terminator      join(5521..5541,1..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 tccttgcgtt ctctaatcct ttcgtcgtgt atattgtaaa aactgctgaa
     2281 cgagtattga taaaagatat cttggccact aaggggcaga tccccttctg gtgtgataga
     2341 caaccccagg agcatagata acaccaactt gtggtggagg gtcatcgaat tggaattgtc
     2401 tgttggcaac agtagaacag atgctgccct tgctatctgt caaaatgccg ctcttcaaag
     2461 ttactttcaa agcgctgtca ctgctacgac agcttttttt ttagaaacag cagcaatgcc
     2521 ttgacatgta acgtaagaaa agaaaaaaga gatggcagaa gaaatactaa gcgataacgg
     2581 caatgtagag gtgctttttt tatcggaata aatagagaag tcagtaacag tgattgctgt
     2641 ggctccctct ttaatcgtat cggatcctgt ttagcttgcc tcgtccccgc cgggtcaccc
     2701 ggccagcgac atggaggccc agaataccct ccttgacagt cttgacgtgc gcagctcagg
     2761 ggcatgatgt gactgtcgcc cgtacattta gcccatacat ccccatgtat aatcatttgc
     2821 atccatacat tttgatggcc gcacggcgcg aagcaaaaat tacggctcct cgctgcagac
     2881 ctgcgagcag ggaaacgctc ccctcacaga cgcgttgaat tgtccccacg ccgcgcccct
     2941 gtagagaaat ataaaaggtt aggatttgcc actgaggttc ttctttcata tacttccttt
     3001 taaaatcttg ctaggataca gttctcacat cacatccgaa cataaacaac catgggtagg
     3061 agggcttttg tagaaagaaa tacgaacgaa acgaaaatca gcgttgccat cgctttggac
     3121 aaagctccct tacctgaaga gtcgaatttt attgatgaac ttataacttc caagcatgca
     3181 aaccaaaagg gagaacaagt aatccaagta gacacgggaa ttggattctt ggatcacatg
     3241 tatcatgcac tggctaaaca tgcaggctgg agcttacgac tttactcaag aggtgattta
     3301 atcatcgatg atcatcacac tgcagaagat actgctattg cacttggtat tgcattcaag
     3361 caggctatgg gtaactttgc cggcgttaaa agatttggac atgcttattg tccacttgac
     3421 gaagcccttt ctagaagcgt agttgacttg tcgggacggc cctatgctgt tatcgatttg
     3481 ggattaaagc gtgaaaaggt tggggaattg tcctgtgaaa tgatccctca cttactatat
     3541 tccttttcgg tagcagctgg aattactttg catgttacct gcttatatgg tagtaatgac
     3601 catcatcgtg ctgaaagcgc ttttaaatct ctggctgttg ccatgcgcgc ggctactagt
     3661 cttactggaa gttctgaagt cccaagcacg aagggagtgt tgtaaagagt actgacaata
     3721 aaaagattct tgttttcaag aacttgtcat ttgtatagtt tttttatatt gtagttgttc
     3781 tattttaatc aaatgttagc gtgatttata ttttttttcg cctcgacatc atctgcccag
     3841 atgcgaagtt aagtgcgcag aaagtaatat catgcgtcaa tcgtatgtga atgctggtcg
     3901 ctatactgct gtcgattcga tactaacgcc gccatccagg ctagcaataa cttcgtataa
     3961 tgtatgctat acgaagttat cagtagagac cgcctggctc tagtagcgat ctacactagc
     4021 actatcagcg ttattaagca ccggtggagt gacgaccttc agcacgttcg tactgttcaa
     4081 cgatggtgta gtcttcgttg tgggaggtga tgtccagttt gatgtcggtt ttgtaagcac
     4141 ccggcagctg aaccggtttt ttagccatgt aggtggtttt aacttcagcg tcgtagtgac
     4201 caccgtcttt cagtttcaga cgcattttga tttcaccttt cagagcaccg tcttccgggt
     4261 acatacgttc ggtggaagct tcccaaccca tggttttttt ctgcataacc ggaccgtcgg
     4321 acgggaagtt ggtaccacgc agtttaactt tgtagatgaa ctcaccgtct tgcagggagg
     4381 agtcctgggt aacggtaaca acaccaccgt cttcgaagtt cataacacgt tcccatttga
     4441 aaccttccgg gaaggacagt ttcaggtagt ccgggatgtc agccgggtgt ttaacgtaag
     4501 ctttggaacc gtactggaac tgcggggaca ggatgtccca agcgaacggc agcggaccac
     4561 ctttggtaac tttcagttta gcggtctggg taccttcgta cggacgacct tcaccttcac
     4621 cttcgatttc gaactcgtga ccgttaacgg aaccttccat acgaactttg aaacgcatga
     4681 actctttgat aacgtcttcg gaggaagcca tctagtattt ctcctctttc tctagtatgt
     4741 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
     4801 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
     4861 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     4921 aggcggtttg cgtattgggt ctcattttat aacttcgtat aatgtatgct atacgaagtt
     4981 atactagtgc atgctttgga cagtgatacc gaagaaggaa gtaaattcag tagcgccgca
     5041 tgaggtagta gcagaaggag ttaaggttga ccatggctca cttggcatat actgttgggc
     5101 agaaacagta tagtgaaact gttatcaaac caatatattt gggctacatg gctttccttt
     5161 cccgaaaaag acaccattta gtaattatgt tgtactttta ctctctgaac ttctgtgaat
     5221 ataaaaaata tactaaataa aaggatgcaa cttccttaaa gagggcaaga tagcgacata
     5281 aagtggaaaa gtgttagcaa cattgcgcag gcatcgatta atatatgcat atatactttt
     5341 tacgtagtct acgttaaatt gtcaatagat caagcagaat cggtagctct gttaagcgat
     5401 taagcccagt ggctttttgt cgcaccgttt aggttcgcgt tagcaagatg gaaaaaaaag
     5461 aacgcgaatc atgtattgac gtatgacgta ttaggcagat gggatacgcg gccgctgcag
     5521 tccggcaaaa aagggcaagg t
//
