LOCUS       sequence-172993-        4882 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tag S. pombe genes in C terminal with mEGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4882)
  AUTHORS   Akamatsu M, Berro J, Pu KM, Tebbs IR, Pollard TD
  TITLE     Cytokinetic nodes in fission yeast arise from two distinct types of 
            nodes that merge during interphase.
  JOURNAL   J Cell Biol. 2014 Mar 17;204(6):977-88. doi: 10.1083/jcb.201307174.
  PUBMED    24637325
REFERENCE   2  (bases 1 to 4882)
  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: "10.1083/jcb"; 
            journalName: "J Cell Biol"; date: "2014-03-17- 17"; volume: "204"; 
            issue: "6"; pages: "977-88"
COMMENT     Sequence Label: pFA6a-mEGFP-KanMX6
FEATURES             Location/Qualifiers
     source          1..4882
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             63..773
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLSPDNHYLSTQSKLSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     CDS             63..773
                     /codon_start=1
                     /product="green fluorescent protein"
                     /label=GFP
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLSPDNHYLSTQSKLSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     primer_bind     complement(90..118)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     713..733
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      797..984
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(852..876)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            1059..2415
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        1059..1402
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1403..2212
                     /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(1470..1489)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      2218..2415
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2519..2538)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2520..2538)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2625..2642)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2796..3384)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2876..2895)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3555..4346,4347..4415))
                     /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     4178..4197
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4416..4520)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4588..4606
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4644..4666)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4766..4785
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        join(4866..4882,1..2)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     join(4866..4882,1)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 gaacgcggcc gccagctgaa gcttcgtacg ctgcaggtcg acggatcccc gggttaatta
       61 acagtaaagg agaagaactt ttcactggag ttgtcccaat tcttgttgaa ttagatggtg
      121 acgttaatgg gcacaaattt tctgtcagtg gagagggtga aggtgatgca acatacggaa
      181 aacttaccct taaatttatt tgcactactg gaaaactacc tgttccatgg ccaacacttg
      241 tcactactct gacttatggt gttcaatgct tttcaagata cccagatcat atgaaacagc
      301 atgacttttt caagagtgcc atgcccgaag gttatgtaca ggaaagaact atatttttca
      361 aagatgacgg gaactacaag acacgtgctg aagtcaagtt tgaaggtgat acccttgtta
      421 atagaatcga gttaaaaggt attgatttta aagaagatgg aaacattctt ggacacaaat
      481 tggaatacaa ctataactca cacaatgtat acatcatggc agacaaacaa aagaatggaa
      541 tcaaagttaa cttcaaaatt agacacaaca ttgaagatgg aagcgttcaa ctagcagacc
      601 attatcaaca aaatactcca attggcgatg gccctgtcct ttcaccagac aaccattacc
      661 tgtccacaca atctaagctt tcgaaagatc ccaacgaaaa gagagaccac atggtccttc
      721 ttgagtttgt aacagctgct gggattacac atggcatgga tgaactatac aaatagggcg
      781 cgccacttct aaataagcga atttcttatg atttatgatt tttattatta aataagttat
      841 aaaaaaaata agtgtataca aattttaaag tgactcttag gttttaaaac gaaaattctt
      901 attcttgagt aactctttcc tgtaggtcag gttgctttct caggtatagt atgaggtcgc
      961 tcttattgac cacacctcta ccggcagatc cgctagggat aacagggtaa tatagatctg
     1021 tttagcttgc ctcgtccccg ccgggtcacc cggccagcga catggaggcc cagaataccc
     1081 tccttgacag tcttgacgtg cgcagctcag gggcatgatg tgactgtcgc ccgtacattt
     1141 agcccataca tccccatgta taatcatttg catccataca ttttgatggc cgcacggcgc
     1201 gaagcaaaaa ttacggctcc tcgctgcaga cctgcgagca gggaaacgct cccctcacag
     1261 acgcgttgaa ttgtccccac gccgcgcccc tgtagagaaa tataaaaggt taggatttgc
     1321 cactgaggtt cttctttcat atacttcctt ttaaaatctt gctaggatac agttctcaca
     1381 tcacatccga acataaacaa ccatgggtaa ggaaaagact cacgtttcga ggccgcgatt
     1441 aaattccaac atggatgctg atttatatgg gtataaatgg gctcgcgata atgtcgggca
     1501 atcaggtgcg acaatctatc gattgtatgg gaagcccgat gcgccagagt tgtttctgaa
     1561 acatggcaaa ggtagcgttg ccaatgatgt tacagatgag atggtcagac taaactggct
     1621 gacggaattt atgcctcttc cgaccatcaa gcattttatc cgtactcctg atgatgcatg
     1681 gttactcacc actgcgatcc ccggcaaaac agcattccag gtattagaag aatatcctga
     1741 ttcaggtgaa aatattgttg atgcgctggc agtgttcctg cgccggttgc attcgattcc
     1801 tgtttgtaat tgtcctttta acagcgatcg cgtatttcgt ctcgctcagg cgcaatcacg
     1861 aatgaataac ggtttggttg atgcgagtga ttttgatgac gagcgtaatg gctggcctgt
     1921 tgaacaagtc tggaaagaaa tgcataagct tttgccattc tcaccggatt cagtcgtcac
     1981 tcatggtgat ttctcacttg ataaccttat ttttgacgag gggaaattaa taggttgtat
     2041 tgatgttgga cgagtcggaa tcgcagaccg ataccaggat cttgccatcc tatggaactg
     2101 cctcggtgag ttttctcctt cattacagaa acggcttttt caaaaatatg gtattgataa
     2161 tcctgatatg aataaattgc agtttcattt gatgctcgat gagtttttct aatcagtact
     2221 gacaataaaa agattcttgt tttcaagaac ttgtcatttg tatagttttt ttatattgta
     2281 gttgttctat tttaatcaaa tgttagcgtg atttatattt tttttcgcct cgacatcatc
     2341 tgcccagatg cgaagttaag tgcgcagaaa gtaatatcat gcgtcaatcg tatgtgaatg
     2401 ctggtcgcta tactgctgtc gattcgatac taacgccgcc atccagttta aacgagctcg
     2461 aattcatcga tgatatcaga tccactagtg gcctatgcgg ccgcggatct gccggtctcc
     2521 ctatagtgag tcgtattaat ttcgataagc caggttaacc tgcattaatg aatcggccaa
     2581 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     2641 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     2701 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     2761 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     2821 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     2881 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     2941 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     3001 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     3061 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     3121 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     3181 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     3241 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     3301 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     3361 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     3421 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     3481 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     3541 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     3601 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     3661 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     3721 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     3781 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     3841 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     3901 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     3961 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     4021 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     4081 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     4141 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     4201 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     4261 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     4321 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     4381 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     4441 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     4501 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc
     4561 attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg
     4621 cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct
     4681 tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc
     4741 gggtgtcggg gctggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat
     4801 atggacatat tgtcgttaga acgcggctac aattaataca taaccttatg tatcatacac
     4861 atacgattta ggtgacacta ta
//
