LOCUS       sequence-148971-        7417 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pCfB2801 is a vector for multiple integrations at sites sharing 
            homology with Ty1Cons2. The selective marker is Kl.LEU2. 
            Additionally it contains a PTEF1-GFP as reporter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7417)
  AUTHORS   Maury J, Germann SM, Baallal Jacobsen SA, Jensen NB, Kildegaard KR, 
            Herrgard MJ, Schneider K, Koza A, Forster J, Nielsen J, Borodina I
  TITLE     EasyCloneMulti: A Set of Vectors for Simultaneous and Multiple 
            Genomic Integrations in Saccharomyces cerevisiae.
  JOURNAL   PLoS One. 2016 Mar 2;11(3):e0150394. doi: 
            10.1371/journal.pone.0150394. eCollection 2016.
  PUBMED    26934490
REFERENCE   2  (bases 1 to 7417)
  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: "PLoS One.";
            date: "2016-03-2"; volume: "11(3):e0150394. doi"; pages: " 
            10.1371/journal.pone.0150394. eCollection 2016"
COMMENT     Sequence Label: pCfB2801
FEATURES             Location/Qualifiers
     source          1..7417
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      6..195
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     protein_bind    243..276
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    2593..2626
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(2669..2688)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2670..2688)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2698..2714)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2707..2729)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     protein_bind    complement(2988..3021)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     primer_bind     complement(3491..3507)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3606..3623)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3777..4365)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3857..3876)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4489..5280,5281..5349))
                     /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     5112..5131
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5350..5441)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5483..5502
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5508..5527
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     protein_bind    5734..5767
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     terminator      complement(6051..6216)
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     6137..6161
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     promoter        6265..6671
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     CDS             6693..7406
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Nager et al., 2011)"
                     /label=superfolder GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATIGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     KYKTRAVVKFEGDTLVNRIELKGTDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFTVRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQTVLSKDPNEKRDHMVLHE
                     YVNAAGITHGMDELYK"
     primer_bind     complement(6723..6751)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
ORIGIN
        1 cattcatccg ctctaaccga aaaggaagga gttagacaac ctgaagtcta ggtccctatt
       61 tattttttta tagttatgtt agtattaaga acgttattta tatttcaaat ttttcttttt
      121 tttctgtaca gacgcgtgta cgcatgtaac attatactga aaaccttgct tgagaaggtt
      181 ttgggacgct cgaagatcgc gtcagctgaa gcttcgtacg ctgcaggtcg acaaccctta
      241 atataacttc gtataatgta tgctatacga agttattagg tctagagatc ccaatacaac
      301 agatcacgtg atctttttgt aagatgaagt tgaagtagtg ttgcaccgtg ccaatgcagg
      361 tggctattag attaaatatg tgatttgttc tattaagttt cctgtataat tacaaatgaa
      421 taacgaaatg agacaaagaa gagaaccaat ttttacaagc agccaagatt tccttgacag
      481 ccttggcgat agcatcgcca acctcagtgg tagagttaga accaccaagg tcaccggttc
      541 tgacacctgc atccaagaca tttctaacag cttcttcaag agccctacct tcttcaacca
      601 aatccaagga taacttcaac atcatagctg cagataagat ggtagcaatt gggttaacct
      661 tgtttgctgg taaatctggg gcagaaccat gacatggttc gtacaaaccg aatgccttgt
      721 tagtgtcagg tagggaagct agagatgcag aaggtaataa acccaaagaa cctggaataa
      781 cagaggcttc atcggagata atatcaccaa acatgttgtt ggtaataaca acaccgttta
      841 gcttagttgg tgatttaacc aaaatcatag cagcagagtc gatcaattgg tgctgaacag
      901 ttaattgtgg gaactcagtc ttgatggttt cttcaacagt ctttctccac aatctggaag
      961 aggcaagcac gttagccttg tcaagtgacc agattggtaa tggtgggttt tgttgcaatg
     1021 ccaagaaagc agccattctt gtaattcttt gaacttcagg aacactgtat ttctcagagt
     1081 cccaagcaac tccgtcacct tcatcttctt ttctttcacc aaagtagata ccaccaacca
     1141 attctctaac gacgacgaaa tcggtaccct ttgcatattc aggcttcaaa ggagaaagat
     1201 ctagtaaaga atcagaagca aagttacatg gtcttaagtt ggcgtataga cccaattcct
     1261 ttctgatctt caatagacct tgttctggtc taactgcgcc cgtaccccat tttggaccac
     1321 caacagcacc tagtaagaca gcatcggctt tcttagaggc ttctagagct tcatctggta
     1381 aaggagtgcc agtggcatcg atggcagcac ccccgatcaa gtgatgttgg aaattgaact
     1441 taatttctgg acggacttca gcaatggcat tcaagacctt aatagcttcg tcagtaactt
     1501 ctttaccgac gtgatcaccc ggtaggacaa cgatattctt agacatggta taatctgtgt
     1561 agtgtgggat actttttact tcttcaaata ggtatcaaca ataaaaataa atcgaagcaa
     1621 atgtaggaat gcgttaaagc agatgtactt acttagagta catagatata tttatataat
     1681 tcaatatata aaagtatatg aagcatctga tgttgaacct gtcatgactc taaactggaa
     1741 tgggtagtta tgcttctgaa ggttttccgt tagaaaatga gtgcagggat caggcccgga
     1801 accggtttta gcctgaaagg aaaagaaaaa aatgcggtga ttactagcac gtgactgcgc
     1861 tgaattggaa tccaccacaa gtacagcgaa tggcaagagg gaaggggaat atattgtgcc
     1921 tggcggtagt ctgctatatc aaatgttaaa cataactaaa tgcagtagtg aatagagtaa
     1981 tgtatttcta catatgtggg tagggccaat gggagcccga tgtctattgt acagatattt
     2041 tccttttata gttaatcaat tcaacttgga aatcattgag tcgatatcta cttggataac
     2101 tggagccgtg gcagcgtttg aaatttgacg agtctccttg ttttgttcgc gcattttcag
     2161 tttattcaaa cgggagatgg caccgaatcc actgacttct ctgttcaatt tgaatggtaa
     2221 taggaagaaa gtacatttcg agaaattttc gcccaacgtc gaaatctcat ggatgatatc
     2281 ctcgatacaa atgacacctt catcacctaa tttctcttca acgatattgt tgtcattcaa
     2341 gattatctct tttggttctt tatcttctgg cctttgccac attattctag atctcttttg
     2401 gattaaagaa cggatagatg caagtgaagg ttgtccgata acaatatatg gtgatacaag
     2461 tttcaacaat gggtacacgt ccttggtaag tttgatgaaa acaccagtgt tcaactcaac
     2521 aagtctcaaa agagataaca ctttgtaagc ctttgctggg atcttcacag cgagctctcg
     2581 agaaccctta atataacttc gtataatgta tgctatacga agttattagg tgatatcaga
     2641 tccactagtg gcctatgcac ccaattcgcc ctatagtgag tcgtattacg cgcgctcact
     2701 ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc gttacccctg caggacagac
     2761 gtcattagtg ctgaggcatt aattgatcat aaaacggaat gatgaataat atttatagaa
     2821 ttgtgtagaa ttgcagattc ccttttatgg attcctaaat cctcgaggag aacttctagt
     2881 atattctgta tacctaatat tatagccttt atcaacaatg gaatcccaac aattatctca
     2941 aaattcacat atttctcaag atctgcggcc gcactcagac ctgaagtgaa gttcctatac
     3001 tttctagaga ataggaactt ctatagtgag tcgaataagg gcgacacaaa atttattcta
     3061 aatgcataat aaatactgat aacatcttat agtttgtatt atattttgta ttatcgttga
     3121 catgtataat tttgatatca aaaactgatt ttccctttat tattttcgag atttattttc
     3181 ttaattctct ttaacaaact agaaatattg tatatacaaa aaatcataaa taatagatga
     3241 atagtttaat tataggtgtt catcaatcga aaaagcaacg tatcttattt aaagtgcgtt
     3301 gcttttttct catttataag gttaaataat tctcatatat caagcaaagt gacaggcgcc
     3361 cttaaatatt ctgacaaatg ctctttccct aaactccccc cataaaaaaa cccgccgaag
     3421 cgggttttta cgttatttgc ggattaacga ttactcgtta tcagaaccgc ccagggggcc
     3481 cgagcttaag actggccgtc gttttacaac acagaaagag tttgtagaaa cgcaaaaagg
     3541 ccatccgtca ggggccttct gcttagtttg atgcctggca gttccctact ctcgccttcc
     3601 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
     3661 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     3721 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     3781 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     3841 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     3901 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     3961 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     4021 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     4081 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     4141 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtgggctaac
     4201 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     4261 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     4321 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     4381 ttttctacgg ggtctgacgc tcagtggaac gacgcgcgcg taactcacgt taagggattt
     4441 tggtcatgag cttgcgccgt cccgtcaagt cagcgtaatg ctctgctttt accaatgctt
     4501 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     4561 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gcgctgcgat
     4621 gataccgcga gaaccacgct caccggctcc ggatttatca gcaataaacc agccagccgg
     4681 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     4741 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     4801 cgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     4861 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     4921 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     4981 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     5041 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     5101 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     5161 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     5221 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     5281 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     5341 aatactcata ttcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     5401 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggtca gtgttacaac
     5461 caattaacca attctgaaca ttatcgcgag cccatttata cctgaatatg gctcataaca
     5521 ccccttgttt gcctggcggc agtagcgcgg tggtcccacc tgaccccatg ccgaactcag
     5581 aagtgaaacg ccgtagcgcc gatggtagtg tggggactcc ccatgcgaga gtagggaact
     5641 gccaggcatc aaataaaacg aaaggctcag tcgaaagact gggcctttcg cccgggctaa
     5701 ttatggggtg tcgcccttat tcgactctat agtgaagttc ctattctcta gaaagtatag
     5761 gaacttctga agtggggatt taaagtcggt gtccgcgctg agggtttaat ggcgcgccgc
     5821 ggccgcccgc ggtgttggaa taaaaatcaa ctatcatcta ctaactagta tttacgttac
     5881 tagtatatta tcatatacgg tgttagaaga tgacgcaaat gatgagaaat agtcatctaa
     5941 attagtggaa gctgaaacgc aaggattgat aatgtaatag gatcaatgaa tattaacata
     6001 gctagcatta agtcctcagc gagcacgctg cttcatggaa tgcgtgcgat gagcgacctc
     6061 atgctatacc tgagaaagca acctgaccta caggaaagag ttactcaaga ataagaattt
     6121 tcgttttaaa acctaagagt cactttaaaa tttgtataca cttatttttt ttataactta
     6181 tttaataata aaaatcataa atcataagaa attcgcttat ttagaagtgt caacaacgta
     6241 tctaccaacg gaatgcgtgc gatgcacaca ccatagcttc aaaatgtttc tactcctttt
     6301 ttactcttcc agattttctc ggactccgcg catcgccgta ccacttcaaa acacccaagc
     6361 acagcatact aaatttcccc tctttcttcc tctagggtgt cgttaattac ccgtactaaa
     6421 ggtttggaaa agaaaaaaga gaccgcctcg tttctttttc ttcgtcgaaa aaggcaataa
     6481 aaatttttat cacgtttctt tttcttgaaa attttttttt ttgatttttt tctctttcga
     6541 tgacctccca ttgatattta agttaataaa cggtcttcaa tttctcaagt ttcagtttca
     6601 tttttcttgt tctattacaa ctttttttac ttcttgctca ttagaaagaa agcatagcaa
     6661 tctaatctaa gttttaatta caaatctgtc atatgagcaa aggagaagaa cttttcactg
     6721 gagttgtccc aattcttgtt gaattagatg gtgatgttaa tgggcacaaa ttttctgtca
     6781 gaggagaggg tgaaggtgat gctacaatcg gaaaactcac ccttaaattt atttgcacta
     6841 ctggaaaact acctgttcca tggccaacac ttgtcactac tctgacctat ggtgttcaat
     6901 gcttttcccg ttatccggat cacatgaaaa ggcatgactt tttcaagagt gccatgcccg
     6961 aaggttatgt acaggaacgc actatatctt tcaaagatga cgggaaatac aagacgcgtg
     7021 ctgtagtcaa gtttgaaggt gatacccttg ttaatcgtat cgagttaaag ggtactgatt
     7081 ttaaagaaga tggaaacatt ctcggacaca aactcgagta caactttaac tcacacaatg
     7141 tatacatcac ggcagacaaa caaaagaatg gaatcaaagc taacttcaca gttcgccaca
     7201 acgttgaaga tggttccgtt caactagcag accattatca acaaaatact ccaattggcg
     7261 atggccctgt ccttttacca gacaaccatt acctgtcgac acaaactgtc ctttcgaaag
     7321 atcccaacga aaagcgtgac cacatggtcc ttcatgagta cgtaaatgct gctgggatta
     7381 cacatggcat ggatgagctc tacaaatgaa tcgcgtg
//
