LOCUS       sequence-203303-        5103 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Yeast gene targeting.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5103)
  AUTHORS   Wu JQ, Kuhn JR, Kovar DR, Pollard TD
  TITLE     Spatial and temporal pathway for assembly and constriction of the 
            contractile ring in fission yeast cytokinesis.
  JOURNAL   Dev Cell. 2003 Nov;5(5):723-34.
  PUBMED    14602073
REFERENCE   2  (bases 1 to 5103)
  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: "Dev Cell.";
            date: "2003-11"; volume: "5(5)"; pages: "723-34"
COMMENT     Sequence Label: pFA6a-mYFP-hphMX6
FEATURES             Location/Qualifiers
     source          1..5103
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        20..38
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     20..37
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             99..812
                     /codon_start=1
                     /product="Aequorea victoria green fluorescent protein"
                     /label=GFP
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTFGYGLKCFARYPDHMKRHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     primer_bind     complement(126..154)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     749..769
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      833..1020
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(888..912)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            1095..2670
                     /label=hphMX6
                     /note="yeast selectable marker conferring hygromycin 
                     resistance"
     promoter        1095..1438
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1439..2467
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLMHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     terminator      2473..2670
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2776..2795)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2777..2795)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2882..2899)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3053..3641)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3133..3152)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3812..4603,4604..4672))
                     /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     4435..4454
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4673..4777)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4845..4863
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4901..4923)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5023..5042
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 tatgtatcat acacatacga tttaggtgac actatagaac gcggccgcca gctgaagctt
       61 cgtacgctgc aggtcgacgg atccccgggt taattaacag taaaggagaa gaacttttca
      121 ctggagttgt cccaattctt gttgaattag atggtgatgt taatgggcac aaattttctg
      181 tcagtggaga gggtgaaggt gatgcaacat acggaaaact tacccttaaa tttatttgca
      241 ctactggaaa actacctgtt ccatggccaa cacttgtcac tactttcggt tatggtctaa
      301 aatgctttgc tagataccca gatcatatga aacggcatga ctttttcaag agtgccatgc
      361 ccgaaggtta tgtacaggaa agaactatat ttttcaaaga tgacgggaac tacaagacac
      421 gtgctgaagt caagtttgaa ggtgataccc ttgttaatag aatcgagtta aaaggtattg
      481 attttaaaga agatggaaac attcttggac acaaattgga atacaactat aactcacaca
      541 atgtatacat catggcagac aaacaaaaga atggaatcaa agttaacttc aaaattagac
      601 acaacattga agatggaagc gttcaactag cagaccatta tcaacaaaat actccaattg
      661 gcgatggccc tgtcctttta ccagacaacc attacctgtc ctatcaatct aagctttcga
      721 aagatcccaa cgaaaagaga gaccacatgg tccttcttga gtttgtaaca gctgctggga
      781 ttacacatgg catggatgaa ctatacaaat agggcgcgcc acttctaaat aagcgaattt
      841 cttatgattt atgattttta ttattaaata agttataaaa aaaataagtg tatacaaatt
      901 ttaaagtgac tcttaggttt taaaacgaaa attcttattc ttgagtaact ctttcctgta
      961 ggtcaggttg ctttctcagg tatagtatga ggtcgctctt attgaccaca cctctaccgg
     1021 cagatccgct agggataaca gggtaatata gatctgttta gcttgcctcg tccccgccgg
     1081 gtcacccggc cagcgacatg gaggcccaga ataccctcct tgacagtctt gacgtgcgca
     1141 gctcaggggc atgatgtgac tgtcgcccgt acatttagcc catacatccc catgtataat
     1201 catttgcatc catacatttt gatggccgca cggcgcgaag caaaaattac ggctcctcgc
     1261 tgcggacctg cgagcaggga aacgctcccc tcacagacgc gttgaattgt ccccacgccg
     1321 cgcccctgta gagaaatata aaaggttagg atttgccact gaggttcttc tttcatatac
     1381 ttccttttaa aatcttgcta ggatacagtt ctcacatcac atccgaacat aaacaaccat
     1441 gggtaaaaag cctgaactca ccgcgacgtc tgtcgagaag tttctgatcg aaaagttcga
     1501 cagcgtctcc gacctgatgc agctctcgga gggcgaagaa tctcgtgctt tcagcttcga
     1561 tgtaggaggg cgtggatatg tcctgcgggt aaatagctgc gccgatggtt tctacaaaga
     1621 tcgttatgtt tatcggcact ttgcatcggc cgcgctcccg attccggaag tgcttgacat
     1681 tggggaattc agcgagagcc tgacctattg catctcccgc cgtgcacagg gtgtcacgtt
     1741 gcaagacctg cctgaaaccg aactgcccgc tgttctgcag ccggtcgcgg aggccatgga
     1801 tgcgatcgct gcggccgatc ttagccagac gagcgggttc ggcccattcg gaccgcaagg
     1861 aatcggtcaa tacactacat ggcgtgattt catatgcgcg attgctgatc cccatgtgta
     1921 tcactggcaa actgtgatgg acgacaccgt cagtgcgtcc gtcgcgcagg ctctcgatga
     1981 gctgatgctt tgggccgagg actgccccga agtccggcac ctcatgcacg cggatttcgg
     2041 ctccaacaat gtcctgacgg acaatggccg cataacagcg gtcattgact ggagcgaggc
     2101 gatgttcggg gattcccaat acgaggtcgc caacatcttc ttctggaggc cgtggttggc
     2161 ttgtatggag cagcagacgc gctacttcga gcggaggcat ccggagcttg caggatcgcc
     2221 gcggctccgg gcgtatatgc tccgcattgg tcttgaccaa ctctatcaga gcttggttga
     2281 cggcaatttc gatgatgcag cttgggcgca gggtcgatgc gacgcaatcg tccgatccgg
     2341 agccgggact gtcgggcgta cacaaatcgc ccgcagaagc gcggccgtct ggaccgatgg
     2401 ctgtgtagaa gtactcgccg atagtggaaa ccgacgcccc agcactcgtc cgagggcaaa
     2461 ggaataatca gtactgacag taaaaagatt cttgttttca agaacttgtc atttgtatag
     2521 tttttttata ttgtagttgt tctattttaa tcaaatgtta gcgtgattta tatttttttt
     2581 cgcctcgaca tcatctgccc agatgcgaag ttaagtgcgc agaaagtaat atcatgcgtc
     2641 aatcgtatgt gaatgctggt cgctatactg ctgtcgattc gatactaacg ccgccatcca
     2701 gtgtttaaac gagctcgaat tcatcgatga tatcagatcc actagtggcc tatgcggccg
     2761 cggatctgcc ggtctcccta tagtgagtcg tattaatttc gataagccag gttaacctgc
     2821 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     2881 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     2941 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     3001 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     3061 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     3121 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     3181 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     3241 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     3301 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     3361 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     3421 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     3481 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     3541 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     3601 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     3661 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     3721 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     3781 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     3841 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     3901 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     3961 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     4021 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     4081 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     4141 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     4201 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     4261 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     4321 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     4381 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     4441 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     4501 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     4561 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     4621 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     4681 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
     4741 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     4801 ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga
     4861 ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc
     4921 cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg
     4981 cgtcagcggg tgttggcggg tgtcggggct ggcttaacta tgcggcatca gagcagattg
     5041 tactgagagt gcaccatatg gacatattgt cgttagaacg cggctacaat taatacataa
     5101 cct
//
