LOCUS       sequence-479482-        4660 bp    DNA     circular SYN 23-JUL-2025
DEFINITION  Tag S. pombe genes at 3' end (C-terminal end of protein) with 
            circularly permuted superfolder GFP, codon-optimized for S. pombe.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4660)
  AUTHORS   Kamenz J, Mihaljev T, Kubis A, Legewie S, Hauf S
  TITLE     Robust Ordering of Anaphase Events by Adaptive Thresholds and 
            Competing Degradation Pathways.
  JOURNAL   Mol Cell. 2015 Nov 5;60(3):446-59. doi: 
            10.1016/j.molcel.2015.09.022. Epub 2015 Oct 29.
  PUBMED    26527280
REFERENCE   2  (bases 1 to 4660)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 23, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.molcel.2015.09.022"; journalName: "Mol Cell"; date: 
            "2015-11-5- 5"; volume: "60"; issue: "3"; pages: "446-59"
COMMENT     Sequence Label: pFA6a-psfGFPcp-natMX6
FEATURES             Location/Qualifiers
     source          1..4660
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      810..997
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(865..889)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            1072..2191
                     /label=natMX6
                     /note="yeast selectable marker conferring nourseothricin 
                     resistance"
     promoter        1072..1415
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1416..1988
                     /codon_start=1
                     /gene="Streptomyces noursei nat1"
                     /product="nourseothricin acetyltransferase"
                     /label=NrsR
                     /note="confers resistance to nourseothricin"
                     /translation="MGTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGF
                     TLREVPVDPPLTKVFPDDESDDESDDGEDGDPDSRTFVAYGDDGDLAGFVVVSYSGWNR
                     RLTVEDIEVAPEHRGHGVGRALMGLATEFARERGAGHLWLEVTNVNAPAIHAYRRMGFT
                     LCGLDTALYDGTASDGEQALYMSMPCP"
     terminator      1994..2191
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2295..2314)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2296..2314)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2401..2418)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2572..3160)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2652..2671)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3331..4122,4123..4191))
                     /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     3954..3973
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4192..4296)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4364..4382
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4420..4442)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4542..4561
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        4642..4660
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     4642..4659
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 acgcggccgc cagctgaagc ttcgtacgct gcaggtcgac ggatccccgg gttaattaac
       61 gaagatggtt ctgttcaact tgcagatcat taccaacaaa acactccaat tggagatggt
      121 ccagttcttc ttcctgataa ccattatctt tctacacaat cagttttgtc taaggatcct
      181 aacgaaaaga gagatcatat ggttttatta gaatttgtta ctgctgctgg tattacacat
      241 ggtatggatg aactttacaa gggaggtggc tctatgcgta agggtgaaga attgtttact
      301 ggagttgttc ctattttagt tgaattggat ggagatgtta atggtcataa gttttctgtt
      361 cgtggtgaag gagaaggaga tgctactaac ggaaaattga cacttaagtt tatttgtact
      421 acaggtaaac ttcctgttcc atggcctact cttgttacta cattgacata tggtgttcaa
      481 tgctttgctc gttacccaga tcatatgaag caacatgatt tctttaagtc agcaatgcct
      541 gaaggatacg ttcaagaacg tactatttct tttaaagatg atggtactta caagacaaga
      601 gctgaggtta agtttgaagg agatactttg gttaaccgaa ttgaacttaa gggtattgat
      661 tttaaggaag atggaaacat tcttggtcat aagcttgaat acaactttaa ctcacataac
      721 gtttacatta ctgctgataa gcaaaagaat ggaattaaag caaactttaa gattcgtcat
      781 aacgtttagg gcgcgccact tctaaataag cgaatttctt atgatttatg atttttatta
      841 ttaaataagt tataaaaaaa ataagtgtat acaaatttta aagtgactct taggttttaa
      901 aacgaaaatt cttattcttg agtaactctt tcctgtaggt caggttgctt tctcaggtat
      961 agtatgaggt cgctcttatt gaccacacct ctaccggcag atccgctagg gataacaggg
     1021 taatatagat ctgtttagct tgcctcgtcc ccgccgggtc acccggccag cgacatggag
     1081 gcccagaata ccctccttga cagtcttgac gtgcgcagct caggggcatg atgtgactgt
     1141 cgcccgtaca tttagcccat acatccccat gtataatcat ttgcatccat acattttgat
     1201 ggccgcacgg cgcgaagcaa aaattacggc tcctcgctgc agacctgcga gcagggaaac
     1261 gctcccctca cagacgcgtt gaattgtccc cacgccgcgc ccctgtagag aaatataaaa
     1321 ggttaggatt tgccactgag gttcttcttt catatacttc ctttttaaat cttgctagga
     1381 tacagttctc acatcacatc cgaacataaa caaccatggg taccactctt gacgacacgg
     1441 cttaccggta ccgcaccagt gtcccggggg acgccgaggc catcgaggca ctggatgggt
     1501 ccttcaccac cgacaccgtc ttccgcgtca ccgccaccgg ggacggcttc accctgcggg
     1561 aggtgccggt ggacccgccc ctgaccaagg tgttccccga cgacgaatcg gacgacgaat
     1621 cggacgacgg ggaggacggc gacccggact ctcggacgtt cgtcgcgtac ggggacgacg
     1681 gcgacctggc gggcttcgtg gtcgtctcgt actccggctg gaaccgccgg ctgaccgtcg
     1741 aggacatcga ggtcgccccg gagcaccggg ggcacggggt cgggcgcgcg ttgatggggc
     1801 tcgcgacgga gttcgcccgc gagcggggtg ccgggcacct ctggctggag gtcaccaacg
     1861 tcaacgcacc ggcgatccac gcgtaccggc ggatggggtt caccctctgc ggcctggaca
     1921 ccgccctgta cgacggcacc gcctcggacg gcgagcaggc gctctacatg agcatgccct
     1981 gcccctaatc agtactgaca ataaaaagat tcttgttttc aagaacttgt catttgtata
     2041 gtttttttat attgtagttg ttctatttta atcaaatgtt agcgtgattt atattttttt
     2101 tcgcctcgac atcatctgcc cagatgcgaa gttaagtgcg cagaaagtaa tatcatgcgt
     2161 caatcgtatg tgaatgctgg tcgctatact gctgtcgatt cgatactaac gccgccatcc
     2221 agtttaaacg agctcgaatt catcgatgat atcagatcca ctagtggcct atgcggccgc
     2281 ggatctgccg gtctccctat agtgagtcgt attaatttcg ataagccagg ttaacctgca
     2341 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     2401 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     2461 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     2521 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     2581 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     2641 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     2701 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     2761 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     2821 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     2881 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     2941 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     3001 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     3061 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     3121 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     3181 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     3241 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     3301 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     3361 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     3421 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
     3481 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     3541 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     3601 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     3661 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     3721 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     3781 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     3841 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     3901 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     3961 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     4021 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     4081 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     4141 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     4201 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     4261 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     4321 tgacgtctaa gaaaccatta ttatcatgac attaacctat aaaaataggc gtatcacgag
     4381 gccctttcgt ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc
     4441 ggagacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc
     4501 gtcagcgggt gttggcgggt gtcggggctg gcttaactat gcggcatcag agcagattgt
     4561 actgagagtg caccatatgg acatattgtc gttagaacgc ggctacaatt aatacataac
     4621 cttatgtatc atacacatac gatttaggtg acactataga
//
