LOCUS       sequence-47080-e        4899 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 4899)
  AUTHORS   Young CL, Raden DL, Caplan JL, Czymmek KJ, Robinson AS
  TITLE     Cassette series designed for live-cell imaging of proteins and 
            high-resolution techniques in yeast.
  JOURNAL   Yeast. 2012 Mar;29(3-4):119-36. doi: 10.1002/yea.2895. Epub 2012 Apr
            4.
  PUBMED    22473760
REFERENCE   2  (bases 1 to 4899)
  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."; 
            date: "2012-03"; volume: "29(3-4):119-36. doi"; pages: " 
            10.1002/yea.2895. Epub 2012 Apr 4"
COMMENT     Sequence Label: pCY 3010-02
FEATURES             Location/Qualifiers
     source          1..4899
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             87..797
                     /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
                     ICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIKAN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITLGMDELYK"
     CDS             87..797
                     /codon_start=1
                     /product="green fluorescent protein"
                     /label=GFP
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIKAN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITLGMDELYK"
     primer_bind     complement(114..142)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     737..757
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     gene            854..2429
                     /label=hphMX6
                     /note="yeast selectable marker conferring hygromycin 
                     resistance"
     promoter        854..1197
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1198..2226
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             1198..2226
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     terminator      2232..2429
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2536..2555)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2537..2555)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2642..2659)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2813..3401)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2893..2912)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3572..4363,4364..4432))
                     /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     4195..4214
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4433..4537)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4605..4623
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4661..4683)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4783..4802
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        join(4883..4899,1..2)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     join(4883..4899,1)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 gaacgcggcc gccagctgaa gcttcgtacg ctgcaggtcg acggatcccc gggcttaaga
       61 tcggtgacgg tgctggttta attaacagta aaggagaaga acttttcact ggagttgtcc
      121 caattcttgt tgaattagat ggtgatgtta atgggcacaa attttctgtc agtggagagg
      181 gtgaaggtga tgcaacatac ggaaaactta cccttaaatt tatttgcact actggaaaac
      241 tacctgttcc atggccaaca cttgtcacta ctttgacttg gggtgttcaa tgctttgcta
      301 gatacccaga tcatatgaaa cagcatgact ttttcaagag tgccatgccc gaaggttatg
      361 tacaggaaag aactatattt ttcaaagatg acgggaacta caagacacgt gctgaagtca
      421 agtttgaagg tgataccctt gttaatagaa tcgagttaaa aggtattgat tttaaagaag
      481 atggaaacat tcttggacac aaattggaat acaacgctat ttcagataat gtatacatca
      541 ctgcagacaa acaaaagaat ggaatcaaag ctaatttcaa aattagacac aacattgaag
      601 atggaagcgt tcaactagca gaccattatc aacaaaatac tccaattggc gatggccctg
      661 tccttttacc agacaaccat tacctgtcca cacaatctgc cctttcgaaa gatcccaacg
      721 aaaagagaga ccacatggtc cttcttgagt ttgtaacagc tgctgggatt acactaggca
      781 tggatgaact atacaaatag ggcgcgccag atctgtttag cttgcctcgt ccccgccggg
      841 tcacccggcc agcgacatgg aggcccagaa taccctcctt gacagtcttg acgtgcgcag
      901 ctcaggggca tgatgtgact gtcgcccgta catttagccc atacatcccc atgtataatc
      961 atttgcatcc atacattttg atggccgcac ggcgcgaagc aaaaattacg gctcctcgct
     1021 gcagacctgc gagcagggaa acgctcccct cacagacgcg ttgaattgtc cccacgccgc
     1081 gcccctgtag agaaatataa aaggttagga tttgccactg aggttcttct ttcatatact
     1141 tccttttaaa atcttgctag gatacagttc tcacatcaca tccgaacata aacaaccatg
     1201 ggtaaaaagc ctgaactcac cgcgacgtct gtcgagaagt ttctgatcga aaagttcgac
     1261 agcgtctccg acctgatgca gctctcggag ggcgaagaat ctcgtgcttt cagcttcgat
     1321 gtaggagggc gtggatatgt cctgcgggta aatagctgcg ccgatggttt ctacaaagat
     1381 cgttatgttt atcggcactt tgcatcggcc gcgctcccga ttccggaagt gcttgacatt
     1441 ggggaattca gcgagagcct gacctattgc atctcccgcc gtgcacaggg tgtcacgttg
     1501 caagacctgc ctgaaaccga actgcccgct gttctgcagc cggtcgcgga ggccatggat
     1561 gcgatcgctg cggccgatct tagccagacg agcgggttcg gcccattcgg accgcaagga
     1621 atcggtcaat acactacatg gcgtgatttc atatgcgcga ttgctgatcc ccatgtgtat
     1681 cactggcaaa ctgtgatgga cgacaccgtc agtgcgtccg tcgcgcaggc tctcgatgag
     1741 ctgatgcttt gggccgagga ctgccccgaa gtccggcacc tcgtgcacgc ggatttcggc
     1801 tccaacaatg tcctgacgga caatggccgc ataacagcgg tcattgactg gagcgaggcg
     1861 atgttcgggg attcccaata cgaggtcgcc aacatcttct tctggaggcc gtggttggct
     1921 tgtatggagc agcagacgcg ctacttcgag cggaggcatc cggagcttgc aggatcgccg
     1981 cggctccggg cgtatatgct ccgcattggt cttgaccaac tctatcagag cttggttgac
     2041 ggcaatttcg atgatgcagc ttgggcgcag ggtcgatgcg acgcaatcgt ccgatccgga
     2101 gccgggactg tcgggcgtac acaaatcgcc cgcagaagcg cggccgtctg gaccgatggc
     2161 tgtgtagaag tactcgccga tagtggaaac cgacgcccca gcactcgtcc gagggcaaag
     2221 gaataatcag tactgacaat aaaaagattc ttgttttcaa gaacttgtca tttgtatagt
     2281 ttttttatat tgtagttgtt ctattttaat caaatgttag cgtgatttat attttttttc
     2341 gcctcgacat catctgccca gatgcgaagt taagtgcgca gaaagtaata tcatgcgtca
     2401 atcgtatgtg aatgctggtc gctatactgc tgtcgattcg atactaacgc cgccatccag
     2461 tgtcgaaaac gagctcgaat tcatcgatga tatcagatcc actagtggcc tatgcggccg
     2521 cggatctgcc ggtctcccta tagtgagtcg tattaatttc gataagccag gttaacctgc
     2581 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     2641 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     2701 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     2761 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     2821 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     2881 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     2941 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     3001 tttctcaatg ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     3061 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     3121 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     3181 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     3241 gctacactag aaggacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     3301 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     3361 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     3421 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     3481 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     3541 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     3601 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     3661 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     3721 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     3781 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     3841 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     3901 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     3961 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     4021 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     4081 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     4141 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     4201 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     4261 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     4321 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     4381 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     4441 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
     4501 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     4561 ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga
     4621 ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc
     4681 cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg
     4741 cgtcagcggg tgttggcggg tgtcggggct ggcttaacta tgcggcatca gagcagattg
     4801 tactgagagt gcaccatatg gacatattgt cgttagaacg cggctacaat taatacataa
     4861 ccttatgtat catacacata cgatttaggt gacactata
//
