LOCUS       sequence-204209-        8115 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Yeast mating-based Split Ubiquitin Assay, prey vector for in vivo 
            cloning.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8115)
  AUTHORS   Obrdlik P, El-Bakkoury M, Hamacher T, Cappellaro C, Vilarino C, 
            Fleischer C, Ellerbrok H, Kamuzinzi R, Ledent V, Blaudez D, Sanders 
            D, Revuelta JL, Boles E, Andre B, Frommer WB
  TITLE     K+ channel interactions detected by a genetic system optimized for 
            systematic studies of membrane protein interactions.
  JOURNAL   Proc Natl Acad Sci U S A. 2004 Aug 17;101(33):12242-7. Epub 2004 Aug
            6.
  PUBMED    15299147
REFERENCE   2  (bases 1 to 8115)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2004-08-17"; volume: "101(33)"; pages: 
            "12242-7. Epub 2004 Aug 6"
COMMENT     Sequence Label: pXNgate22-1HA
FEATURES             Location/Qualifiers
     source          1..8115
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..17
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        40..744
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 promoter
                     /note="promoter for alcohol dehydrogenase 1"
     protein_bind    754..778
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     gene            865..2221
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        865..1208
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1209..2018
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(1276..1295)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      2024..2221
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     protein_bind    complement(2275..2299)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             2445..2471
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     terminator      2496..2683
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2551..2575)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     primer_bind     complement(2707..2723)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2707..2723)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2720..2742)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2731..2747
                     /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(2755..2761,2762..2779,2780..2785))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2800..2821
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2938..2955)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3109..3697)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3189..3208)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3868..4659,4660..4728))
                     /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     4491..4510
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4729..4833)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4860..6202
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    complement(5823..5870)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     primer_bind     6243..6261
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6299..6321)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6421..6440
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        6461..6742
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     CDS             6743..7417
                     /codon_start=1
                     /gene="S. cerevisiae TRP1"
                     /product="phosphoribosylanthranilate isomerase, required 
                     for tryptophan biosynthesis"
                     /label=TRP1
                     /note="yeast auxotrophic marker"
                     /translation="MSVINFTGSSGPLVKVCGLQSTEAAECALDSDADLLGIICVPNRK
                     RTIDPVIARKISSLVKAYKNSSGTPKYLVGVFRNQPKEDVLALVNDYGIDIVQLHGDES
                     WQEYQEFLGLPVIKRLVFPKDCNILLSAASQKPHSFIPLFDSEAGGTGELLDWNSISDW
                     VGRQESPESLHFMLAGGLTPENVGDALRLNGVIGVDVSGGVETNGVKDSNKIANFVKNA
                     KK"
     rep_origin      complement(7520..7975)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7657..7678)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7869..7888
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     join(8101..8115,1..8)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     join(8115,1..17)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
ORIGIN
        1 gtaaaacgac ggccagtgcc aagctttcgc gagctcgata tccttttgtt gtttccgggt
       61 gtacaatatg gacttcctct tttctggcaa ccaaacccat acatcgggat tcctataata
      121 ccttcgttgg tctccctaac atgtaggtgg cggaggggag atatacaata gaacagatac
      181 cagacaagac ataatgggct aaacaagact acaccaatta cactgcctca ttgatggtgg
      241 tacataacga actaatactg tagccctaga cttgatagcc atcatcatat cgaagtttca
      301 ctaccctttt tccatttgcc atctattgaa gtaataatag gcgcatgcaa cttcttttct
      361 ttttttttct tttctctctc ccccgttgtt gtctcaccat atccgcaatg acaaaaaaat
      421 gatggaagac actaaaggaa aaaattaacg acaaagacag caccaacaga tgtcgttgtt
      481 ccagagctga tgaggggtat ctcgaagcac acgaaacttt ttccttcctt cattcacgca
      541 cactactctc taatgagcaa cggtatacgg ccttccttcc agttacttga atttgaaata
      601 aaaaaaagtt tgctgtcttg ctatcaagta taaatagacc tgcaattatt aatcttttgt
      661 ttcctcgtca ttgttctcgt tccctttctt ccttgtttct ttttctgcac aatatttcaa
      721 gctataccaa gcatacaatc aactcatacg tgaacaagtt tgtacaaaaa agcaggctct
      781 ccaaccacca tgaattctat acgaagttat taggtctaga gatctgttta gcttgcctcg
      841 tccccgccgg gtcacccggc cagcgacatg gaggcccaga ataccctcct tgacagtctt
      901 gacgtgcgca gctcaggggc atgatgtgac tgtcgcccgt acatttagcc catacatccc
      961 catgtataat catttgcatc catacatttt gatggccgca cggcgcgaag caaaaattac
     1021 ggctcctcgc tgcagacctg cgagcaggga aacgctcccc tcacagacgc gttgaattgt
     1081 ccccacgccg cgcccctgta gagaaatata aaaggttagg atttgccact gaggttcttc
     1141 tttcatatac ttccttttaa aatcttgcta ggatacagtt ctcacatcac atccgaacat
     1201 aaacaaccat gggtaaggaa aagactcacg tttcgaggcc gcgattaaat tccaacatgg
     1261 atgctgattt atatgggtat aaatgggctc gcgataatgt cgggcaatca ggtgcgacaa
     1321 tctatcgatt gtatgggaag cccgatgcgc cagagttgtt tctgaaacat ggcaaaggta
     1381 gcgttgccaa tgatgttaca gatgagatgg tcagactaaa ctggctgacg gaatttatgc
     1441 ctcttccgac catcaagcat tttatccgta ctcctgatga tgcatggtta ctcaccactg
     1501 cgatccccgg caaaacagca ttccaggtat tagaagaata tcctgattca ggtgaaaata
     1561 ttgttgatgc gctggcagtg ttcctgcgcc ggttgcattc gattcctgtt tgtaattgtc
     1621 cttttaacag cgatcgcgta tttcgtctcg ctcaggcgca atcacgaatg aataacggtt
     1681 tggttgatgc gagtgatttt gatgacgagc gtaatggctg gcctgttgaa caagtctgga
     1741 aagaaatgca taagcttttg ccattctcac cggattcagt cgtcactcat ggtgatttct
     1801 cacttgataa ccttattttt gacgagggga aattaatagg ttgtattgat gttggacgag
     1861 tcggaatcgc agaccgatac caggatcttg ccatcctatg gaactgcctc ggtgagtttt
     1921 ctccttcatt acagaaacgg ctttttcaaa aatatggtat tgataatcct gatatgaata
     1981 aattgcagtt tcatttgatg ctcgatgagt ttttctaatc agtactgaca ataaaaagat
     2041 tcttgttttc aagaacttgt catttgtata gtttttttat attgtagttg ttctatttta
     2101 atcaaatgtt agcgtgattt atattttttt tcgcctcgac atcatctgcc cagatgcgaa
     2161 gttaagtgcg cagaaagtaa tatcatgcgt caatcgtatg tgaatgctgg tcgctatact
     2221 gctgtcgatt cgatactaac gccgccatcc agtgtcgaaa acgagctccc gggtacccag
     2281 ctttcttgta caaagtggtt ggtggtggcg gatccggtgg aggtggatca atgcagattt
     2341 tcgtcaagac tttgaccggt aaaaccggaa cattggaagt tgaatcttcc gataccatcg
     2401 acaacgttaa gtcgaaaatt caagacaagg aaggaatccc tggttaccca tacgacgttc
     2461 cagattatgc ttagctgcag ccaagctaat tccgggcgaa tttcttatga tttatgattt
     2521 ttattattaa ataagttata aaaaaaataa gtgtatacaa attttaaagt gactcttagg
     2581 ttttaaaacg aaaattctta ttcttgagta actctttcct gtaggtcagg ttgctttctc
     2641 aggtatagca tgaggtcgct cttattgacc acacctctac cggcatgcaa gcttggcgta
     2701 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     2761 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     2821 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     2881 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     2941 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     3001 ggctgtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     3061 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     3121 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     3181 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     3241 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     3301 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     3361 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     3421 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     3481 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     3541 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     3601 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     3661 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     3721 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     3781 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     3841 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     3901 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     3961 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     4021 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     4081 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     4141 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     4201 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     4261 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     4321 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     4381 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     4441 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     4501 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     4561 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     4621 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     4681 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     4741 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     4801 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
     4861 acgaagcatc tgtgcttcat tttgtagaac aaaaatgcaa cgcgagagcg ctaatttttc
     4921 aaacaaagaa tctgagctgc atttttacag aacagaaatg caacgcgaaa gcgctatttt
     4981 accaacgaag aatctgtgct tcatttttgt aaaacaaaaa tgcaacgcga gagcgctaat
     5041 ttttcaaaca aagaatctga gctgcatttt tacagaacag aaatgcaacg cgagagcgct
     5101 attttaccaa caaagaatct atacttcttt tttgttctac aaaaatgcat cccgagagcg
     5161 ctatttttct aacaaagcat cttagattac tttttttctc ctttgtgcgc tctataatgc
     5221 agtctcttga taactttttg cactgtaggt ccgttaaggt tagaagaagg ctactttggt
     5281 gtctattttc tcttccataa aaaaagcctg actccacttc ccgcgtttac tgattactag
     5341 cgaagctgcg ggtgcatttt ttcaagataa aggcatcccc gattatattc tataccgatg
     5401 tggattgcgc atactttgtg aacagaaagt gatagcgttg atgattcttc attggtcaga
     5461 aaattatgaa cggtttcttc tattttgtct ctatatacta cgtataggaa atgtttacat
     5521 tttcgtattg ttttcgattc actctatgaa tagttcttac tacaattttt ttgtctaaag
     5581 agtaatacta gagataaaca taaaaaatgt agaggtcgag tttagatgca agttcaagga
     5641 gcgaaaggtg gatgggtagg ttatataggg atatagcaca gagatatata gcaaagagat
     5701 acttttgagc aatgtttgtg gaagcggtat tcgcaatatt ttagtagctc gttacagtcc
     5761 ggtgcgtttt tggttttttg aaagtgcgtc ttcagagcgc ttttggtttt caaaagcgct
     5821 ctgaagttcc tatactttct agagaatagg aacttcggaa taggaacttc aaagcgtttc
     5881 cgaaaacgag cgcttccgaa aatgcaacgc gagctgcgca catacagctc actgttcacg
     5941 tcgcacctat atctgcgtgt tgcctgtata tatatataca tgagaagaac ggcatagtgc
     6001 gtgtttatgc ttaaatgcgt acttatatgc gtctatttat gtaggatgaa aggtagtcta
     6061 gtacctcctg tgatattatc ccattccatg cggggtatcg tatgcttcct tcagcactac
     6121 cctttagctg ttctatatgc tgccactcct caattggatt agtctcatcc ttcaatgcta
     6181 tcatttcctt tgatattgga tcatactaag aaaccattat tatcatgaca ttaacctata
     6241 aaaataggcg tatcacgagg ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc
     6301 tctgacacat gcagctcccg gagacggtca cagcttgtct gtaagcggat gccgggagca
     6361 gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg tcggggctgg cttaactatg
     6421 cggcatcaga gcagattgta ctgagagtgc accatagatc aacgacatta ctatatatat
     6481 aatataggaa gcatttaata gaacagcatc gtaatatatg tgtactttgc agttatgacg
     6541 ccagatggca gtagtggaag atattcttta ttgaaaaata gcttgtcacc ttacgtacaa
     6601 tcttgatccg gagcttttct ttttttgccg attaagaatt aattcggtcg aaaaaagaaa
     6661 aggagagggc caagagggag ggcattggtg actattgagc acgtgagtat acgtgattaa
     6721 gcacacaaag gcagcttgga gtatgtctgt tattaatttc acaggtagtt ctggtccatt
     6781 ggtgaaagtt tgcggcttgc agagcacaga ggccgcagaa tgtgctctag attccgatgc
     6841 tgacttgctg ggtattatat gtgtgcccaa tagaaagaga acaattgacc cggttattgc
     6901 aaggaaaatt tcaagtcttg taaaagcata taaaaatagt tcaggcactc cgaaatactt
     6961 ggttggcgtg tttcgtaatc aacctaagga ggatgttttg gctctggtca atgattacgg
     7021 cattgatatc gtccaactgc atggagatga gtcgtggcaa gaataccaag agttcctcgg
     7081 tttgccagtt attaaaagac tcgtatttcc aaaagactgc aacatactac tcagtgcagc
     7141 ttcacagaaa cctcattcgt ttattccctt gtttgattca gaagcaggtg ggacaggtga
     7201 acttttggat tggaactcga tttctgactg ggttggaagg caagagagcc ccgaaagctt
     7261 acattttatg ttagctggtg gactgacgcc agaaaatgtt ggtgatgcgc ttagattaaa
     7321 tggcgttatt ggtgttgatg taagcggagg tgtggagaca aatggtgtaa aagactctaa
     7381 caaaatagca aatttcgtca aaaatgctaa gaaataggtt attactgagt agtatttatt
     7441 taagtattgt ttgtgcactt gccgatctat gcggtgtgaa ataccgcaca gatgcgtaag
     7501 gagaaaatac cgcatcagga aattgtaagc gttaatattt tgttaaaatt cgcgttaaat
     7561 ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa
     7621 tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta
     7681 ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca
     7741 ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat
     7801 cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg
     7861 agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc
     7921 acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtccatt
     7981 cgccattcag gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct tcgctattac
     8041 gccagctggc gaaaggggga tgtgctgcaa ggcgattaag ttgggtaacg ccagggtttt
     8101 cccagtcacg acgtt
//
