LOCUS       sequence-203271-        4888 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 4888)
  AUTHORS   Liu Y, Lee IJ, Sun M, Lower CA, Runge KW, Ma J, Wu JQ
  TITLE     Roles of the novel coiled-coil protein Rng10 in septum formation 
            during fission yeast cytokinesis.
  JOURNAL   Mol Biol Cell. 2016 Aug 15;27(16):2528-41. doi: 
            10.1091/mbc.E16-03-0156. Epub 2016 Jul 6.
  PUBMED    27385337
REFERENCE   2  (bases 1 to 4888)
  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: 
            "10.1091/mbc.E16-03-0156"; journalName: "Mol Biol Cell"; date: 
            "2016-08-15- 15"; volume: "27"; issue: "16"; pages: "2528-41"
COMMENT     Sequence Label: pFA6a-mMaple3-kanMX6
FEATURES             Location/Qualifiers
     source          1..4888
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        15..33
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     15..32
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     terminator      834..1021
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(889..913)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            1096..2452
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        1096..1439
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1440..2249
                     /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(1507..1526)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      2255..2452
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2556..2575)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2557..2575)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2662..2679)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2833..3421)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2913..2932)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3592..4383,4384..4452))
                     /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     4215..4234
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4453..4557)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4625..4643
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4681..4703)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4803..4822
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 atcatacaca tacgatttag gtgacactat agaacgcggc cgccagctga agcttcgtac
       61 gctgcaggtc gacggatccc cgggttaatt aacggtagtg gagttagcaa gggcgaggag
      121 accatcatga gcgtgatcaa gccggacatg aagatcaagc tgcgcatgga gggcaacgtg
      181 aacggccatg cctttgtgat cgagggcgag ggcagcggta agccgtttga gggcatccag
      241 accatcgacc tggaggttaa ggaaggcgca ccgctgccgt ttgcctacga catcctgacc
      301 accgcattcc actacggcaa ccgcgtgttc accaagtacc cgcgcaagat cccggactac
      361 ttcaagcaga gcttcccgga gggctacagt tgggaacgca gcatgaccta cgaggacggc
      421 ggtatctgca acgccaccaa cgacatcacc atggaagaag atagcttcat caacaagatc
      481 cacttcaagg gcacaaactt cccgccgaat ggtccggtta tgcagaagcg caccgttggc
      541 tgggaggtga gcaccgagaa gatgtatgtg cgcgacggtg tgctgaaggg cgacgtgaag
      601 atgaagctgc tgctgaaggg tggcagccac taccgctgcg acttccgcac cacctacaaa
      661 gttaagcaaa aggcagtgaa gttaccgaag gcccacttcg tggaccaccg catcgaaatc
      721 ctgagccacg acaaggacta taacaaagtg aagctgtacg agcacgccgt ggcccgtaac
      781 agcaccgaca gcatggatga gctgtacaaa taaggcgcgc cacttctaaa taagcgaatt
      841 tcttatgatt tatgattttt attattaaat aagttataaa aaaaataagt gtatacaaat
      901 tttaaagtga ctcttaggtt ttaaaacgaa aattcttatt cttgagtaac tctttcctgt
      961 aggtcaggtt gctttctcag gtatagtatg aggtcgctct tattgaccac acctctaccg
     1021 gcagatccgc tagggataac agggtaatat agatctgttt agcttgcctc gtccccgccg
     1081 ggtcacccgg ccagcgacat ggaggcccag aataccctcc ttgacagtct tgacgtgcgc
     1141 agctcagggg catgatgtga ctgtcgcccg tacatttagc ccatacatcc ccatgtataa
     1201 tcatttgcat ccatacattt tgatggccgc acggcgcgaa gcaaaaatta cggctcctcg
     1261 ctgcagacct gcgagcaggg aaacgctccc ctcacagacg cgttgaattg tccccacgcc
     1321 gcgcccctgt agagaaatat aaaaggttag gatttgccac tgaggttctt ctttcatata
     1381 cttcctttta aaatcttgct aggatacagt tctcacatca catccgaaca taaacaacca
     1441 tgggtaagga aaagactcac gtttcgaggc cgcgattaaa ttccaacatg gatgctgatt
     1501 tatatgggta taaatgggct cgcgataatg tcgggcaatc aggtgcgaca atctatcgat
     1561 tgtatgggaa gcccgatgcg ccagagttgt ttctgaaaca tggcaaaggt agcgttgcca
     1621 atgatgttac agatgagatg gtcagactaa actggctgac ggaatttatg cctcttccga
     1681 ccatcaagca ttttatccgt actcctgatg atgcatggtt actcaccact gcgatccccg
     1741 gcaaaacagc attccaggta ttagaagaat atcctgattc aggtgaaaat attgttgatg
     1801 cgctggcagt gttcctgcgc cggttgcatt cgattcctgt ttgtaattgt ccttttaaca
     1861 gcgatcgcgt atttcgtctc gctcaggcgc aatcacgaat gaataacggt ttggttgatg
     1921 cgagtgattt tgatgacgag cgtaatggct ggcctgttga acaagtctgg aaagaaatgc
     1981 ataagctttt gccattctca ccggattcag tcgtcactca tggtgatttc tcacttgata
     2041 accttatttt tgacgagggg aaattaatag gttgtattga tgttggacga gtcggaatcg
     2101 cagaccgata ccaggatctt gccatcctat ggaactgcct cggtgagttt tctccttcat
     2161 tacagaaacg gctttttcaa aaatatggta ttgataatcc tgatatgaat aaattgcagt
     2221 ttcatttgat gctcgatgag tttttctaat cagtactgac aataaaaaga ttcttgtttt
     2281 caagaacttg tcatttgtat agttttttta tattgtagtt gttctatttt aatcaaatgt
     2341 tagcgtgatt tatatttttt ttcgcctcga catcatctgc ccagatgcga agttaagtgc
     2401 gcagaaagta atatcatgcg tcaatcgtat gtgaatgctg gtcgctatac tgctgtcgat
     2461 tcgatactaa cgccgccatc cagtttaaac gagctcgaat tcatcgatga tatcagatcc
     2521 actagtggcc tatgcggccg cggatctgcc ggtctcccta tagtgagtcg tattaatttc
     2581 gataagccag gttaacctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     2641 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     2701 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     2761 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     2821 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     2881 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     2941 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     3001 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     3061 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     3121 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     3181 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     3241 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     3301 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     3361 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     3421 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     3481 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     3541 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     3601 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     3661 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     3721 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     3781 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     3841 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     3901 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     3961 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     4021 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     4081 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     4141 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     4201 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     4261 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     4321 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     4381 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     4441 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     4501 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     4561 atttccccga aaagtgccac ctgacgtcta agaaaccatt attatcatga cattaaccta
     4621 taaaaatagg cgtatcacga ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa
     4681 cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag
     4741 cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggct ggcttaacta
     4801 tgcggcatca gagcagattg tactgagagt gcaccatatg gacatattgt cgttagaacg
     4861 cggctacaat taatacataa ccttatgt
//
