LOCUS       sequence-149089-        7915 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For integration of Cre recombinase at the yeast HIS3 locus orthe 
            CYC1 terminator.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7915)
  AUTHORS   Terweij M, van Welsem T, van Deventer S, Verzijlbergen KF, 
            Menendez-Benito V, Ontoso D, San-Segundo P, Neefjes J, van Leeuwen F
  TITLE     Recombination-induced tag exchange (RITE) cassette series to monitor
            protein dynamics in Saccharomyces cerevisiae.
  JOURNAL   G3 (Bethesda). 2013 Aug 7;3(8):1261-72. doi: 10.1534/g3.113.006213.
  PUBMED    23708297
REFERENCE   2  (bases 1 to 7915)
  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: "10.1534/g3.113"; 
            journalName: "G3 (Bethesda)"; date: "2013-08-7- 7"; volume: "3"; 
            issue: "8"; pages: "1261-72"
COMMENT     Sequence Label: pTW040
FEATURES             Location/Qualifiers
     source          1..7915
                     /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"
     promoter        317..504
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             505..1167
                     /codon_start=1
                     /gene="S. cerevisiae HIS3"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=HIS3
                     /note="yeast auxotrophic marker"
                     /translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE
                     QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF
                     KEALGAVRGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHF
                     LESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM"
     rep_origin      complement(1427..1882)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1564..1585)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1776..1795
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2027..2043
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        2050..2068
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     2101..2117
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2101..2117
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     promoter        join(2209..2214,2215..2232,2233..2239)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2247..2263
                     /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)."
     primer_bind     2271..2287
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        2308..2326
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        2397..3039
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     2933..2956
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     CDS             3046..4074
                     /codon_start=1
                     /product="site-specific recombinase"
                     /label=Cre
                     /note="Cre is a site-specific recombinase from 
                     bacteriophage P1. Recombination occurs at loxP sequences."
                     /translation="MSNLLTVHQNLPALPVDATSDEVRKNLMDMFRDRQAFSEHTWKML
                     LSVCRSWAAWCKLNNRKWFPAEPEDVRDYLLYLQARGLAVKTIQQHLGQLNMLHRRSGL
                     PRPSDSNAVSLVMRRIRKENVDAGERAKQALAFERTDFDQVRSLMENSDRCQDIRNLAF
                     LGIAYNTLLRIAEIARIRVKDISRTDGGRMLIHIGRTKTLVSTAGVEKALSLGVTKLVE
                     RWISVSGVADDPNNYLFCRVRKNGVAAPSATSQLSTRALEGIFEAAHRLIYGAEDDSGQ
                     RYLAWSGHSARVGAARDMARAGVSIPEIMQAGGWTNVNIVMNYIRNLDSETGAMVVLLE
                     DGD"
     CDS             4087..5022
                     /codon_start=1
                     /product="mutated ligand-binding domain of the human 
                     estrogen receptor (Feil et al., 1997)"
                     /label=ERT2
                     /translation="AGDMRAANLWPSPLMIKRSKENSLALSLTADQMVSALLDAEPPIL
                     YSEYDPTRPFSEASMMGLLTNLADRELVHMINWAKRVPGFVDLTLHDQVHLLECAWLGI
                     LMIGLVWRSMEHPGKLLFAPNLLLDRNQGKCVEGMVEIFDMLLVTSSRFRMMNLQGEEF
                     VCLKSIILLNSGVYTFLSSTLKSLEEKDHIHRVLDKITDTLIHLMAKAGLTLQQQHQRL
                     AQLLLLLSHIRHMSNEGMEHLYSMKCKNVVPLYDLLLEMLDAHRLHAPTSRGGASVEET
                     DQSHLATAGSTSSHSLQKYYITGEAEGFPAT"
     terminator      5199..5446
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(5215..5233)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     promoter        complement(5465..5483)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5493..5509)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5609..5625)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5610..5626)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     promoter        complement(5655..5673)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5694..5710)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    5718..5734
                     /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(5742..5748,5749..5766,5767..5772))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(5925..5942)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6096..6684)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6176..6195)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6855..7646,7647..7715))
                     /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     7478..7497
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7716..7820)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7888..7906
                     /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 accataaatt cccgttttaa gagcttggtg agcgctagga gtcactgcca ggtatcgttt
      241 gaacacggca ttagtcaggg aagtcataac acagtccttt cccgcaattt tctttttcta
      301 ttactcttgg cctcctctag tacactctat atttttttat gcctcggtaa tgattttcat
      361 tttttttttt ccacctagcg gatgactctt tttttttctt agcgattggc attatcacat
      421 aatgaattat acattatata aagtaatgtg atttcttcga agaatatact aaaaaatgag
      481 caggcaagat aaacgaaggc aaagatgaca gagcagaaag ccctagtaaa gcgtattaca
      541 aatgaaacca agattcagat tgcgatctct ttaaagggtg gtcccctagc gatagagcac
      601 tcgatcttcc cagaaaaaga ggcagaagca gtagcagaac aggccacaca atcgcaagtg
      661 attaacgtcc acacaggtat agggtttctg gaccatatga tacatgctct ggccaagcat
      721 tccggctggt cgctaatcgt tgagtgcatt ggtgacttac acatagacga ccatcacacc
      781 actgaagact gcgggattgc tctcggtcaa gcttttaaag aggccctagg ggccgtgcgt
      841 ggagtaaaaa ggtttggatc aggatttgcg cctttggatg aggcactttc cagagcggtg
      901 gtagatcttt cgaacaggcc gtacgcagtt gtcgaacttg gtttgcaaag ggagaaagta
      961 ggagatctct cttgcgagat gatcccgcat tttcttgaaa gctttgcaga ggctagcaga
     1021 attaccctcc acgttgattg tctgcgaggc aagaatgatc atcaccgtag tgagagtgcg
     1081 ttcaaggctc ttgcggttgc cataagagaa gccacctcgc ccaatggtac caacgatgtt
     1141 ccctccacca aaggtgttct tatgtagtga caccgattat ttaaagctgc agcatacgat
     1201 atatatacat gtgtatatat gtatacctat gaatgtcagt aagtatgtat acgaacagta
     1261 tgatactgaa gatgacaagg taatgcatca ttctatacgt gtcattctga acgaggcgcg
     1321 ctttcctttt ttctttttgc tttttctttt tttttctctt gaactcgacg gatctatgcg
     1381 gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggaaat tgtaaacgtt
     1441 aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag
     1501 gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt
     1561 gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga
     1621 aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg
     1681 gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct
     1741 tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc
     1801 gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt
     1861 aatgcgccgc tacagggcgc gtcgcgccat tcgccattca ggctgcgcaa ctgttgggaa
     1921 gggcgatcgg tgcgggcctc ttcgctatta cgccagctgg cgaaaggggg atgtgctgca
     1981 aggcgattaa gttgggtaac gccagggttt tcccagtcac gacgttgtaa aacgacggcc
     2041 agtgaattgt aatacgactc actatagggc gaattggagc tccaccgcgg tggcggccgc
     2101 tctagaacta gtggatcccc cctggcacga caggtttccc gactggaaag cgggcagtga
     2161 gcgcaacgca attaatgtga gttacctcac tcattaggca ccccaggctt tacactttat
     2221 gcttccggct cctatgttgt gtggaattgt gagcggataa caatttcaca caggaaacag
     2281 ctatgaccat gattacgcca agcgcgcaat taaccctcac taaagggaac aaaagctggg
     2341 ggcccggaaa cagctatgac catgattacg ccaagcttgg ctgcaggtcg agtttatcat
     2401 tatcaatact cgccatttca aagaatacgt aaataattaa tagtagtgat tttcctaact
     2461 ttatttagtc aaaaaattag ccttttaatt ctgctgtaac ccgtacatgc caaaataggg
     2521 ggcgggttac acagaatata taacactgat ggtgcttggg tgaacaggtt tattcctggc
     2581 atccactaaa tataatggag cccgcttttt aagctggcat ccagaaaaaa aaagaatccc
     2641 agcaccaaaa tattgttttc ttcaccaacc atcagttcat aggtccattc tcttagcgca
     2701 actacagaga acagggcaca aacaggcaaa aaacgggcac aacctcaatg gagtgatgca
     2761 acctgcctgg agtaaatgat gacacaaggc aattgaccca cgcatgtatc tatctcattt
     2821 tcttacacct tctattacct tctgctctct ctgatttgga aaaagctgaa aaaaaaggtt
     2881 taaaccagtt ccctgaaatt attcccctac ttgactaata agtatataaa gacggtaggt
     2941 attgattgta attctgtaaa tctatttctt aaacttctta aattctactt ttatagttag
     3001 tctttttttt agttttaaaa caccaagaac ttagtttcgt ccggaatgtc caatttactg
     3061 accgtacacc aaaatttgcc tgcattaccg gtcgatgcaa cgagtgatga ggttcgcaag
     3121 aacctgatgg acatgttcag ggatcgccag gcgttttctg agcatacctg gaaaatgctt
     3181 ctgtccgttt gccggtcgtg ggcggcatgg tgcaagttga ataaccggaa atggtttccc
     3241 gcagaacctg aagatgttcg cgattatctt ctatatcttc aggcgcgcgg tctggcagta
     3301 aaaactatcc agcaacattt gggccagcta aacatgcttc atcgtcggtc cgggctgcca
     3361 cgaccaagtg acagcaatgc tgtttcactg gttatgcggc ggatccgaaa agaaaacgtt
     3421 gatgccggtg aacgtgcaaa acaggctcta gcgttcgaac gcactgattt cgaccaggtt
     3481 cgttcactca tggaaaatag cgatcgctgc caggatatac gtaatctggc atttctgggg
     3541 attgcttata acaccctgtt acgtatagcc gaaattgcca ggatcagggt taaagatatc
     3601 tcacgtactg acggtgggag aatgttaatc catattggca gaacgaaaac gctggttagc
     3661 accgcaggtg tagagaaggc acttagcctg ggggtaacta aactggtcga gcgatggatt
     3721 tccgtctctg gtgtagctga tgatccgaat aactacctgt tttgccgggt cagaaaaaat
     3781 ggtgttgccg cgccatctgc caccagccag ctatcaactc gcgccctgga agggattttt
     3841 gaagcagctc atcgactgat ttacggcgct gaggatgact ctggtcagag gtacctggcc
     3901 tggtctggac acagtgcccg tgtcggagcc gcgcgagata tggcccgcgc tggagtttca
     3961 ataccggaga tcatgcaagc tggtggctgg accaatgtaa atattgtcat gaactatatc
     4021 cgtaacctgg atagtgaaac aggggcaatg gtggtcctgc tggaagatgg cgatctcgag
     4081 ccatccgctg gagacatgag agctgccaac ctttggccaa gcccgctcat gatcaaacgc
     4141 tctaaggaga acagcctggc cttgtccctg acggccgacc agatggtcag tgccttgttg
     4201 gatgctgagc cccccatact ctattccgag tatgatccta ccagaccctt cagtgaagct
     4261 tcgatgatgg gcttactgac caacctggca gacagggagc tggttcacat gatcaactgg
     4321 gcgaagaggg tgccaggctt tgtggatttg accctccatg atcaggtcca ccttctagaa
     4381 tgtgcctggc tagggatcct gatgattggt ctcgtctggc gctccatgga acacccgggg
     4441 aagctcctgt ttgctcctaa cttgctcctg gacaggaatc aaggtaaatg tgtggaaggc
     4501 atggtggaga tcttcgacat gctgctggtt acatcatctc ggttccgcat gatgaatctg
     4561 cagggagagg agtttgtgtg cctcaaatct attattttgc ttaattctgg agtatacaca
     4621 tttctgtcca gcaccctgaa gtctctcgag gagaaggacc atatccaccg agtcctggac
     4681 aagatcacag acactttgat ccacctgatg gccaaggcag gcctgaccct gcagcagcag
     4741 caccagcggc tagcccagct cctcctcctc ctctcccaca tcaggcacat gagtaacgaa
     4801 ggcatggagc atctgtacag catgaagtgc aagaacgtgg tacccctcta tgacctgctg
     4861 ctggagatgc tggacgccca ccgcctacat gcgcccacta gtcgtggagg ggcatccgtg
     4921 gaggagacgg accaaagcca cttggccact gcgggctcta cttcatcgca ttccttgcaa
     4981 aagtattaca tcacggggga ggcagagggt ttccctgcca cagtctgagt agaatgcctg
     5041 atattgactc aatatccgtt gcgtttcctg tcaaaagtat gcgtagtgct gaacatttcg
     5101 tgatgaatgc caccgaggaa gaagcacggc gcggttttgc taaagtgatg tctgagtttg
     5161 gcgaactctt gggtaaggtt ggaattgtcg acctcgagtc atgtaattag ttatgtcacg
     5221 cttacattca cgccctcccc ccacatccgc tctaaccgaa aaggaaggag ttagacaacc
     5281 tgaagtctag gtccctattt atttttttat agttatgtta gtattaagaa cgttatttat
     5341 atttcaaatt tttctttttt ttctgtacag acgcgtgtac gcatgtaaca ttatactgaa
     5401 aaccttgctt gagaaggttt tgggacgctc gaaggcttta atttgcggcc ggtacccaat
     5461 tcgccctata gtgagtcgta ttacgcgcgc tcactggccg tcgttttaca acgtcgtgac
     5521 tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagg
     5581 ggctgcagga attcgatatc aagcttatcg ataccgtcga cctcgagggg gggcccggta
     5641 cccagctttt gttcccttta gtgagggtta attccgagct tggcgtaatc atggtcatag
     5701 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     5761 ataaagtgta aagcctgggg tgcctaatga gtgaggtaac tcacattaat tgcgttgcgc
     5821 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     5881 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     5941 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     6001 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     6061 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     6121 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     6181 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     6241 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     6301 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     6361 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     6421 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     6481 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca
     6541 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     6601 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     6661 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     6721 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     6781 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     6841 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     6901 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     6961 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     7021 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     7081 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     7141 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     7201 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     7261 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     7321 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     7381 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     7441 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     7501 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     7561 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     7621 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     7681 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     7741 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     7801 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc
     7861 attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtk
//
