LOCUS       sequence-425142-        8773 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A pHis5 backbone integrating plasmid expressing mECitrine under the 
            inducible promoter 41nmt1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8773)
  AUTHORS   Pan ZQ, Yang YS, Du LL
  TITLE     Pil1 Co-tethering Assay to Detect Protein-Protein Interactions in 
            the Fission Yeast Schizosaccharomyces pombe.
  JOURNAL   Methods Mol Biol. 2025;2862:93-102. doi: 
            10.1007/978-1-0716-4168-2_7.
  PUBMED    39527195
REFERENCE   2  (bases 1 to 8773)
  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: "Methods Mol
            Biol."; date: "2025"; volume: "2862:93-102. doi"; pages: " 
            10.1007/978-1-0716-4168-2_7"
COMMENT     Sequence Label: pDB4884
FEATURES             Location/Qualifiers
     source          1..8773
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     CDS             1188..1838
                     /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="MRRAFVERNTNETKISVAIALDKAPLPEESNFIDELITSKHANQK
                     GEQVIQVDTGIGFLDHMYHALAKHAGWSLRLYSRGDLIIDDHHTAEDTAIALGIAFKQA
                     MGNFAGVKRFGHAYCPLDEALSRSVVDLSGRPYAVIDLGLKREKVGELSCEMIPHLLYS
                     FSVAAGITLHVTCLYGSNDHHRAESAFKSLAVAMRAATSLTGSSEVPSTKGVL"
     rep_origin      complement(2272..2727)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2409..2430)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2621..2640
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2853..2875
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2867..2884
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2868..2884
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2894..2913
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2894..2912
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4133..4161)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     4756..4776
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      4844..5031
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(4899..4923)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            5100..6456
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        5100..5443
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             5444..6253
                     /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(5511..5530)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      6259..6456
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(6513..6533)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(6513..6531)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(6552..6568)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6552..6568)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6565..6587)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6576..6592
                     /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(6600..6606,6607..6624,6625..6630))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6645..6666
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6783..6800)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6954..7542)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7034..7053)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7713..8504,8505..8573))
                     /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     8336..8355
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8574..8678)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     8746..8764
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accataaccg ggatggaacg attgaatact gtgctactct ttatgtaata taacgaattc
      241 aactgttcaa cgtatgtgac tgctctctgc ttctcggtat atatttactt gaaggaatta
      301 atttttcttt gccaatggtc tcttttacca attgcagtct ggcttaaagc aatgaggttg
      361 taattaaatg gcttgagacc tagaacaaac accttttaca atccttgttt actggttttg
      421 atagtaaatt cggttcgctt gtggtgtaca ttatattttt taggctagtc gtgatttttt
      481 gataatatca atattcttag atttgagaca ttcatgcttt ttaatttgat caatcgtttg
      541 tatcttcatt aattagtagg tggtttgata gaacatttag ttgcttgttt gactacaaat
      601 atctgcagaa tggattcggt taataaaaaa aatgtttgag gaattaattt tcgctttgga
      661 gtggaggagc ggccgctcct caattcgatg ttgcagattt tacaagtttt taaaatgtat
      721 ttcattatta ctttttatat gcctaataaa aaagccatag tttaatctat agataacttt
      781 ttttccagtg cactaacgga cgttacattc ccatacaaaa ctgcgtagtt aaagctaagg
      841 aaaagttaat atcatgttaa ttaaatacgc tatttacaat aagacattga actcatttat
      901 atcgttgaat atgaataacc aatttcagcg aatttttaac aaacatcgtt cacctcgttt
      961 aaggatatct tgtgtatggg gtgttgactt gctttatcga ataattaccg tacctgtaat
     1021 tggcttgctg gatatagcgg tagtctaata tctagcaaaa atcttttggg tgaaaaggct
     1081 tgcaatttca cgacaccgaa ctatttgtca ttttttaata aggaagtttt ccataaattc
     1141 ctgtaattct cggttgatct aattgaaaag agtagttttg catcacgatg aggagggctt
     1201 ttgtagaaag aaatacgaac gaaacgaaaa tcagcgttgc catcgctttg gacaaagctc
     1261 ccttacctga agagtcgaat tttattgatg aacttataac ttccaagcat gcaaaccaaa
     1321 agggagaaca agtaatccaa gtagacacgg gaattggatt cttggatcac atgtatcatg
     1381 cactggctaa acatgcaggc tggagcttac gactttactc aagaggtgat ttaatcatcg
     1441 atgatcatca cactgcagaa gatactgcta ttgcacttgg tattgcattc aagcaggcta
     1501 tgggtaactt tgccggcgtt aaaagatttg gacatgctta ttgtccactt gacgaagctc
     1561 tttctagaag cgtagttgac ttgtcgggac ggccctatgc tgttatcgat ttgggattaa
     1621 agcgtgaaaa ggttggggaa ttgtcctgtg aaatgatccc tcacttacta tattcctttt
     1681 cggtagcagc tggaattact ttgcatgtta cctgcttata tggtagtaat gaccatcatc
     1741 gtgctgaaag cgcttttaaa tctctggctg ttgccatgcg cgcggctact agtcttactg
     1801 gaagttctga agtcccaagc acgaagggag tgttgtaaag atgaattgga ttatgtcagg
     1861 aaaagaacga caattttgca tccaaattgt ctaaatttta gagttgcttg aaaacaatag
     1921 aaccttactt gctttataat tacgttaatt agaagcgtta tctcgtgaag gaatatagta
     1981 cgtagccgta taaattgaat tgaatgttca gcttatagaa tagagacact ttgctgttca
     2041 atgcgtcgtc acttaccata ctcactttat tatacgactt taagtataaa ctccgcggtt
     2101 atggtaaaat taatgatgca caaacgtccg attccatatg ggtacactac aattaaatac
     2161 ttttaagtaa cgctgatttc ttatttttta tccttggtgt atttattgtt tataccgcta
     2221 gggcggtgtg aaataccgca cagatgcgta aggagaaaat accgcatcag gaaattgtaa
     2281 gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc
     2341 aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga
     2401 gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag
     2461 ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt
     2521 ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta
     2581 gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag
     2641 cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg
     2701 cgcttaatgc gccgctacag ggcgcgtcca ttcgccattc aggctgcgca actgttggga
     2761 agggcgatcg gtgcgggcct cttcgctatt acgccagctg gcgaaagggg gatgtgctgc
     2821 aaggcgatta agttgggtaa cgccagggtt ttcccagtca cgacgttgta aaacgacggc
     2881 cagtgagcgc gcgtaatacg actcactata gggcgaattg ggtaccgggc ccgatcagaa
     2941 aattatcgcc ataaaagaca gaataagtca tcagcggttg tttcatttcc tatatttttt
     3001 ttttattttt ttatttttta ataagggaaa atttaacgtc taaggataca gaagattgtt
     3061 agcacattaa agtaataaag gcttaagtag taagtgcctt agcatgttat tgtatttcaa
     3121 aggacataat ctaaaataat aacaatatca tttctcacaa gttattcaat tttctttttt
     3181 tttctaataa tatcaagaat gtattatttg tttgacataa gtcaactaat ttatttaata
     3241 tgctggatta atcttgcaga catgtaaatt aacaagtttt agtcaaataa cgttgaagtt
     3301 tcaatgaact caaataattt ctcttttttt ttatataacc ataatctgat ttatattttc
     3361 cgcagggatc aactgaagtt atgacatttg gattggatca cttataacct tggtcgccaa
     3421 ataatacaaa aatcagcgtt ataaaacaaa gaaggttttt gttaagaaat taatcctctt
     3481 tcttgataag aaagttgaac cgaaattgca gatactgata tatgaaaata atacccacaa
     3541 ttttgggaat agcgcaagcc tcaatttaaa caataggtga ggacacatga taatgacctc
     3601 aatgattgtt agaagaaaag agcctcatta caaaatcgaa aaatgaatgg ttgggtacaa
     3661 gtttccaaaa catggtaaag tggactttgc gtatgagacg taaatagaaa aaaacacttg
     3721 ttatatgttt tctagaatta ttgttgtctc tttatggttg gatgatgcaa aatagtaatt
     3781 tcggttagtt gctgtaaaac accacgagac aaatagatat ggatatttat taaatcagga
     3841 aaaacgtaac tctcggctac tggatggttc agtcacccaa cgattactgg ggagagaaaa
     3901 cagggcaaaa gcaaagctta aaggaatccg attgtcattc ggcaatgtgc agcgaaacta
     3961 aaaaccggat aatggacctg ttaatcgaaa cattgaagat aaaggaagag gaatcctggc
     4021 atatcatcaa ttgaataagt tgaattaatt atttcaatct cattctcact ttctgactta
     4081 tagtcgcttt gttaaagcta gcatgagtaa aggagaagaa cttttcactg gagttgtccc
     4141 aattcttgtt gaattagatg gtgatgttaa tgggcacaaa ttttctgtca gtggagaggg
     4201 tgaaggtgat gcaacatacg gaaaacttac ccttaaattt atttgcacta ctggaaaact
     4261 acctgttcca tggccaacac ttgtcactac tttgggttat ggtctaatgt gctttgctag
     4321 atacccagat catatgaaac ggcatgactt tttcaagagt gccatgcccg aaggttatgt
     4381 acaggaaaga actatatttt tcaaagatga cgggaactac aagacacgtg ctgaagtcaa
     4441 gtttgaaggt gatacccttg ttaatagaat cgagttaaaa ggtattgatt ttaaagaaga
     4501 tggaaacatt cttggacaca aattggaata caactataac tcacacaatg tatacatcat
     4561 ggcagacaaa caaaagaatg gaatcaaagt taacttcaaa attagacaca acattgaaga
     4621 tggaagcgtt caactagcag accattatca acaaaatact ccaattggcg atggccctgt
     4681 ccttttacca gacaaccatt acctgtccta tcaatctaag ctttcgaaag atcccaacga
     4741 aaagagagac cacatggtcc ttcttgagtt tgtaacagct gctgggatta cacatggcat
     4801 ggatgaacta tacaaaagat cttaacgcgc cacttctaaa taagcgaatt tcttatgatt
     4861 tatgattttt attattaaat aagttataaa aaaaataagt gtatacaaat tttaaagtga
     4921 ctcttaggtt ttaaaacgaa aattcttatt cttgagtaac tctttcctgt aggtcaggtt
     4981 gctttctcag gtatagtatg aggtcgctct tattgaccac acctctaccg gcagatccgc
     5041 tagggataac agggtaatat gtttagcttg cctcgacccc gccgggtcac ccggccagcg
     5101 acatggaggc ccagaatacc ctccttgaca gtcttgacgt gcgcagctca ggggcatgat
     5161 gtgactgtcg cccgtacatt tagcccatac atccccatgt ataatcattt gcatccatac
     5221 attttgatgg ccgcacggcg cgaagcaaaa attacggctc ctcgctgcag acctgcgagc
     5281 agggaaacgc tcccctcaca gacgcgttga attgtcccca cgccgcgccc ctgtagagaa
     5341 atataaaagg ttaggatttg ccactgaggt tcttctttca tatacttcct tttaaaatct
     5401 tgctaggata cagttctcac atcacatccg aacataaaca accatgggta aggaaaagac
     5461 tcacgtttcg aggccgcgat taaattccaa catggatgct gatttatatg ggtataaatg
     5521 ggctcgcgat aatgtcgggc aatcaggtgc gacaatctat cgattgtatg ggaagcccga
     5581 tgcgccagag ttgtttctga aacatggcaa aggtagcgtt gccaatgatg ttacagatga
     5641 gatggtcaga ctaaactggc tgacggaatt tatgcctctt ccgaccatca agcattttat
     5701 ccgtactcct gatgatgcat ggttactcac cactgcgatc cccggcaaaa cagcattcca
     5761 ggtattagaa gaatatcctg attcaggtga aaatattgtt gatgcgctgg cagtgttcct
     5821 gcgccggttg cattcgattc ctgtttgtaa ttgtcctttt aacagcgatc gcgtatttcg
     5881 tctcgctcag gcgcaatcac gaatgaataa cggtttggtt gatgcgagtg attttgatga
     5941 cgagcgtaat ggctggcctg ttgaacaagt ctggaaagaa atgcataagc ttttgccatt
     6001 ctcaccggat tcagtcgtca ctcatggtga tttctcactt gataacctta tttttgacga
     6061 ggggaaatta ataggttgta ttgatgttgg acgagtcgga atcgcagacc gataccagga
     6121 tcttgccatc ctatggaact gcctcggtga gttttctcct tcattacaga aacggctttt
     6181 tcaaaaatat ggtattgata atcctgatat gaataaattg cagtttcatt tgatgctcga
     6241 tgagtttttc taatcagtac tgacaataaa aagattcttg ttttcaagaa cttgtcattt
     6301 gtatagtttt tttatattgt agttgttcta ttttaatcaa atgttagcgt gatttatatt
     6361 ttttttcgcc tcgacatcat ctgcccagat gcgaagttaa gtgcgcagaa agtaatatca
     6421 tgcgtcaatc gtatgtgaat gctggtcgct atactgctgt cgattcgata ctaacgccgc
     6481 catccagttt aaacgagctc cagcttttgt tccctttagt gagggttaat tgcgcgcttg
     6541 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     6601 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     6661 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     6721 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
     6781 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     6841 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     6901 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     6961 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     7021 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     7081 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     7141 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     7201 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     7261 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     7321 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     7381 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     7441 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     7501 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     7561 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     7621 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     7681 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     7741 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     7801 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     7861 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     7921 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     7981 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     8041 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     8101 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     8161 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     8221 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     8281 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     8341 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     8401 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     8461 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     8521 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     8581 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     8641 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     8701 cctgacgtct aagaaaccat tattatcatg acattaacct ataaaaatag gcgtatcacg
     8761 aggccctttc gtc
//
