LOCUS       sequence-56250-e        5395 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5395)
  AUTHORS   Zilio N, Wehrkamp-Richter S, Boddy MN
  TITLE     A new versatile system for rapid control of gene expression in the 
            fission yeast Schizosaccharomyces pombe.
  JOURNAL   Yeast. 2012 Aug 29. doi: 10.1002/yea.2920.
  PUBMED    22968950
REFERENCE   2  (bases 1 to 5395)
  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: "Yeast. 2012
            Aug 29. doi: 10.1002/yea.2920."
COMMENT     Sequence Label: pFA6a-kanMX-tetO-Pcyc1-GFP
FEATURES             Location/Qualifiers
     source          1..5395
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(1..19)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2..19)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(100..119)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     219..241
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(279..297)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        365..469
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(470..538,539..1330)
                     /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     complement(688..707)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1501..2089
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1990..2009
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2243..2260
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     2347..2366
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2347..2365
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     gene            complement(2470..3826)
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     terminator      complement(2470..2667)
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     CDS             complement(2673..3482)
                     /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     3396..3415
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(3483..3826)
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     protein_bind    join(3909..3927,3928..3950,3951..3969,3970..3992,3993..4011,
                     4012..4034,4035..4053,4054..4076,4077..4095,4096..4118,
                     4119..4137,4138..4160,4161..4179)
                     /label=tetracycline response element
                     /bound_moiety="tetracycline repressor TetR"
                     /note="contains seven copies of the tetracycline operator 
                     tetO"
     CDS             4399..5109
                     /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
                     ICTTGKLPVPWPTLVTTFTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     primer_bind     complement(4426..4454)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     5049..5069
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      5133..5320
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(5188..5212)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
ORIGIN
        1 tctatagtgt cacctaaatc gtatgtgtat gatacataag gttatgtatt aattgtagcc
       61 gcgttctaac gacaatatgt ccatatggtg cactctcagt acaatctgct ctgatgccgc
      121 atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct
      181 gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag
      241 gttttcaccg tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac gcctattttt
      301 ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa
      361 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
      421 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
      481 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
      541 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
      601 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
      661 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
      721 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
      781 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
      841 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
      901 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
      961 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
     1021 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     1081 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     1141 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     1201 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     1261 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     1321 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     1381 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     1441 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     1501 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     1561 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     1621 cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt
     1681 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     1741 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     1801 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     1861 ctacaccgaa ctgagatacc tacagcgtga gcattgagaa agcgccacgc ttcccgaagg
     1921 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     1981 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     2041 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     2101 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     2161 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     2221 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
     2281 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcaggttaa cctggcttat
     2341 cgaaattaat acgactcact atagggagac cggcagatcc gcggccgcat aggccactag
     2401 tggatctgat atcatcgatg aattcgagct cgtttaaact ggatggcggc gttagtatcg
     2461 aatcgacagc agtatagcga ccagcattca catacgattg acgcatgata ttactttctg
     2521 cgcacttaac ttcgcatctg ggcagatgat gtcgaggcga aaaaaaatat aaatcacgct
     2581 aacatttgat taaaatagaa caactacaat ataaaaaaac tatacaaatg acaagttctt
     2641 gaaaacaaga atctttttat tgtcagtact gattagaaaa actcatcgag catcaaatga
     2701 aactgcaatt tattcatatc aggattatca ataccatatt tttgaaaaag ccgtttctgt
     2761 aatgaaggag aaaactcacc gaggcagttc cataggatgg caagatcctg gtatcggtct
     2821 gcgattccga ctcgtccaac atcaatacaa cctattaatt tcccctcgtc aaaaataagg
     2881 ttatcaagtg agaaatcacc atgagtgacg actgaatccg gtgagaatgg caaaagctta
     2941 tgcatttctt tccagacttg ttcaacaggc cagccattac gctcgtcatc aaaatcactc
     3001 gcatcaacca aaccgttatt cattcgtgat tgcgcctgag cgagacgaaa tacgcgatcg
     3061 ctgttaaaag gacaattaca aacaggaatc gaatgcaacc ggcgcaggaa cactgccagc
     3121 gcatcaacaa tattttcacc tgaatcagga tattcttcta atacctggaa tgctgttttg
     3181 ccggggatcg cagtggtgag taaccatgca tcatcaggag tacggataaa atgcttgatg
     3241 gtcggaagag gcataaattc cgtcagccag tttagtctga ccatctcatc tgtaacatca
     3301 ttggcaacgc tacctttgcc atgtttcaga aacaactctg gcgcatcggg cttcccatac
     3361 aatcgataga ttgtcgcacc tgattgcccg acattatcgc gagcccattt atacccatat
     3421 aaatcagcat ccatgttgga atttaatcgc ggcctcgaaa cgtgagtctt ttccttaccc
     3481 atggttgttt atgttcggat gtgatgtgag aactgtatcc tagcaagatt ttaaaaggaa
     3541 gtatatgaaa gaagaacctc agtggcaaat cctaaccttt tatatttctc tacaggggcg
     3601 cggcgtgggg acaattcaac gcgtctgtga ggggagcgtt tccctgctcg caggtctgca
     3661 gcgaggagcc gtaatttttg cttcgcgccg tgcggccatc aaaatgtatg gatgcaaatg
     3721 attatacatg gggatgtatg ggctaaatgt acgggcgaca gtcacatcat gcccctgagc
     3781 tgcgcacgtc aagactgtca aggagggtat tctgggcctc catgtcgctg gccgggtgac
     3841 ccggcgggga cgaggcaagc taaacagatc tgggtaatat agatcaattc ctcgatcgag
     3901 tttaccactc cctatcagtg atagagaaaa gtgaaagtcg agtttaccac tccctatcag
     3961 tgatagagaa aagtgaaagt cgagtttacc actccctatc agtgatagag aaaagtgaaa
     4021 gtcgagttta ccactcccta tcagtgatag agaaaagtga aagtcgagtt taccactccc
     4081 tatcagtgat agagaaaagt gaaagtcgag tttaccactc cctatcagtg atagagaaaa
     4141 gtgaaagtcg agtttaccac tccctatcag tgatagagaa aagtgaaagt cgagctcggt
     4201 accctatggc atgcatgtgc tctgtatgta tataaaactc ttgttttctt cttttctcta
     4261 aatattcttt ccttatacat taggtccttt gtagcataaa ttactatact tctatagaca
     4321 cgcaaacaca aatacacaca ctaaattacc ggatcaattc gggggatccg tttatcatcc
     4381 tggaggatgt taattaacag taaaggagaa gaacttttca ctggagttgt cccaattctt
     4441 gttgaattag atggtgatgt taatgggcac aaattttctg tcagtggaga gggtgaaggt
     4501 gatgcaacat acggaaaact tacccttaaa tttatttgca ctactggaaa actacctgtt
     4561 ccatggccaa cacttgtcac tactttcact tatggtgttc aatgcttttc aagataccca
     4621 gatcatatga aacggcatga ctttttcaag agtgccatgc ccgaaggtta tgtacaggaa
     4681 agaactatat ttttcaaaga tgacgggaac tacaagacac gtgctgaagt caagtttgaa
     4741 ggtgataccc ttgttaatag aatcgagtta aaaggtattg attttaaaga agatggaaac
     4801 attcttggac acaaattgga atacaactat aactcacaca atgtatacat catggcagac
     4861 aaacaaaaga atggaatcaa agttaacttc aaaattagac acaacattga agatggaagc
     4921 gttcaactag cagaccatta tcaacaaaat actccaattg gcgatggccc tgtcctttta
     4981 ccagacaacc attacctgtc cacacaatct gccctttcga aagatcccaa cgaaaagaga
     5041 gaccacatgg tccttcttga gtttgtaaca gctgctggga ttacacatgg catggatgaa
     5101 ctatacaaat agggcgcgcc acttctaaat aagcgaattt cttatgattt atgattttta
     5161 ttattaaata agttataaaa aaaataagtg tatacaaatt ttaaagtgac tcttaggttt
     5221 taaaacgaaa attcttattc ttgagtaact ctttcctgta ggtcaggttg ctttctcagg
     5281 tatagtatga ggtcgctctt attgaccaca cctctaccgg cagatccgct agggataaca
     5341 gggtaatata gatccgtcga cctgcagcgt acgaagcttc agctggcggc cgcgt
//
