LOCUS       sequence-128281-        7987 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Integration of 2GFP tag at Atg2 C terminus, use auxotrophic marker 
            URA3(Candida albicans).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7987)
  AUTHORS   Li D, Song JZ, Shan MH, Li SP, Liu W, Li H, Zhu J, Wang Y, Lin J, 
            Xie Z
  TITLE     A fluorescent tool set for yeast Atg proteins.
  JOURNAL   Autophagy. 2015;11(6):954-60. doi: 10.1080/15548627.2015.1040971.
  PUBMED    25998947
REFERENCE   2  (bases 1 to 7987)
  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: 
            "Autophagy."; date: "2015"; volume: "11(6):954-60. doi"; pages: " 
            10.1080/15548627.2015.1040971"
COMMENT     Sequence Label: LD202
FEATURES             Location/Qualifiers
     source          1..7987
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(30..458)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(140..161)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     352..371
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     585..607
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     599..616
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     677..693
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     677..693
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(727..743)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(728..744)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     CDS             1973..2686
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGIDELYK"
     CDS             2723..3439
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             2723..3436
                     /codon_start=1
                     /product="green fluorescent protein"
                     /label=GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     terminator      3460..3647
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(3515..3539)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     promoter        3722..4065
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     terminator      4884..5081
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(5799..5819)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5799..5817)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5838..5854)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5838..5854)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5851..5873)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5862..5878
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(5886..5892,5893..5910,5911..5916))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5931..5952
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6069..6086)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6240..6828)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6320..6339)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6999..7790,7791..7859))
                     /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     7622..7641
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7860..7964)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
       61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
      121 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
      181 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
      241 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
      301 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
      361 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
      421 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgagc gcgcgtaata cgactcacta tagggcgaat tggagctcca
      661 ccgcggtggc ggccgctcta gaactagtgg atcccccggg ctgcaggaat tcgatatcaa
      721 gcttatcgat accgtcgacc tcgagtttct tctggacctg gaatttatcc ggccgatcga
      781 ctttttgcaa gcagtggaat tagtcatgca attaaatgtt gcgcctttac gccttcacgt
      841 ggatcaagac actttagagt ttctaataag gttcttaggg ttcaaagata aaagatttga
      901 attaattgat gagtatccag atattgtatt tattcaaaaa tttagcacta attcaattaa
      961 gttaagatta gattataagc cgaaaaaagt tgattatgcg ggtttaagat cgggccaaac
     1021 ttctgaattg atgaattttt ttacccttga tggatccaag attattttga aaagcgtagt
     1081 cctgtatgga ttaaatggat tcgatgaatt aaataataaa ctgaaagcca tttggacgcc
     1141 ggatattacc aaaaagcagc tgcctggcgt tttggaagga ctggctcctg tcagatcgtt
     1201 tatggcgatt gggtcagggg tgaagaccct agtcacagta ctaatgtcag aatacaggca
     1261 agaagggcat ttgggaagga gtctgaaaaa gggtggcaat gtcttcttga aaacgacaac
     1321 gggagatttt gtgaaattgg gagttaaatt aacttcaggg acacaggcga tattggagaa
     1381 caccgaggaa ctattcggtg gtgttggttc caatggcagg gtgtacgatg catcgaaatt
     1441 tggctccgct gatggtgcag acagtgatac tgcagctgta ttagacttgg atactttatt
     1501 cgaggaggat cagttagttg gtagtaaata ttccaggata agagatcatg agcccacggc
     1561 agtagtcatc gatatgtctt caccgggaga tcacaacgaa cccacaatag tgagtctgta
     1621 cgcagatcag cccctcgacc tgcccacggg tttgaaggag gcgtatagtt cattagaaaa
     1681 gcatatgcat attgcgtacg atgcggtatg gagggcaaag ggacagatga aggatgataa
     1741 acggggaggg ccgagcgccg cagcagttta tgttgctaga gctgcacctg tggcaattat
     1801 tcggccgtta attggggcca ctgaggcagt gtctaagact ttacagggaa tagctaatca
     1861 agtggataag acgcataacg agcaaatcaa tgataagtac aagtccaatc ggactgattc
     1921 ggcttcgtac gctgcaggtc gacggatcgg tgacggtgct ggtttaatta acatgtctaa
     1981 aggtgaagaa ttattcactg gtgttgtccc aattttggtt gaattagatg gtgatgttaa
     2041 tggtcacaaa ttttctgtct ccggtgaagg tgaaggtgat gctacttacg gtaaattgac
     2101 cttaaaattt atttgtacta ctggtaaatt gccagttcca tggccaacct tagtcactac
     2161 tttaacttat ggtgttcaat gtttttctag atacccagat catatgaaac aacatgactt
     2221 tttcaagtct gccatgccag aaggttatgt tcaagaaaga actatttttt tcaaagatga
     2281 cggtaactac aagaccagag ctgaagtcaa gtttgaaggt gataccttag ttaatagaat
     2341 cgaattaaaa ggtattgatt ttaaagaaga tggtaacatt ttaggtcaca aattggaata
     2401 caactataac tctcacaatg tttacatcat ggctgacaaa caaaagaatg gtatcaaagt
     2461 taacttcaaa attagacaca acattgaaga tggttctgtt caattagctg accattatca
     2521 acaaaatact ccaattggtg atggtccagt cttgttacca gacaaccatt acttatccac
     2581 tcaatctgcc ttatccaaag atccaaacga aaagagagac cacatggtct tgttagaatt
     2641 tgttactgct gctggtatta cccatggtat tgatgaattg tacaaaattc cagttccaac
     2701 gcgtgatcta gagaagccat tcatgtctaa aggtgaagaa ttattcactg gtgttgtccc
     2761 aattttggtt gaattagatg gtgatgttaa tggtcacaaa ttttctgtct ccggtgaagg
     2821 tgaaggtgat gctacttacg gtaaattgac cttaaaattt atttgtacta ctggtaaatt
     2881 gccagttcca tggccaacct tagtcactac tttaacttat ggtgttcaat gtttttctag
     2941 atacccagat catatgaaac aacatgactt tttcaagtct gccatgccag aaggttatgt
     3001 tcaagaaaga actatttttt tcaaagatga cggtaactac aagaccagag ctgaagtcaa
     3061 gtttgaaggt gataccttag ttaatagaat cgaattaaaa ggtattgatt ttaaagaaga
     3121 tggtaacatt ttaggtcaca aattggaata caactataac tctcacaatg tttacatcat
     3181 ggctgacaaa caaaagaatg gtatcaaagt taacttcaaa attagacaca acattgaaga
     3241 tggttctgtt caattagctg accattatca acaaaatact ccaattggtg atggtccagt
     3301 cttgttacca gacaaccatt acttatccac tcaatctgcc ttatccaaag atccaaacga
     3361 aaagagagac cacatggtct tgttagaatt tgttactgct gctggtatta cccatggtat
     3421 ggatgaatta tataaataag gcgcgccact tctaaataag cgaatttctt atgatttatg
     3481 atttttatta ttaaataagt tataaaaaaa ataagtgtat acaaatttta aagtgactct
     3541 taggttttaa aacgaaaatt cttattcttg agtaactctt tcctgtaggt caggttgctt
     3601 tctcaggtat agtatgaggt cgctcttatt gaccacacct ctaccggcag atccgctagg
     3661 gataacaggg taatatagat ctgtttagct tgcctcgtcc ccgccgggtc acccggccag
     3721 cgacatggag gcccagaata ccctccttga cagtcttgac gtgcgcagct caggggcatg
     3781 atgtgactgt cgcccgtaca tttagcccat acatccccat gtataatcat ttgcatccat
     3841 acattttgat ggccgcacgg cgcgaagcaa aaattacggc tcctcgctgc agacctgcga
     3901 gcagggaaac gctcccctca cagacgcgtt gaattgtccc cacgccgcgc ccctgtagag
     3961 aaatataaaa ggttaggatt tgccactgag gttcttcttt catatacttc cttttaaaat
     4021 cttgctagga tacagttctc acatcacatc cgaacataaa caaccatgac agtcaacact
     4081 aagacctata gtgagagagc agaaactcat gcctcaccag tagcacaacg attatttcga
     4141 ttaatggaac tgaagaaaac caatttatgt gcatcaattg atgttgatac cactaaggaa
     4201 ttccttgaat taattgataa attgggtcct tatgtatgct taatcaagac tcatattgat
     4261 ataatcaatg atttttccta tgaatccact attgaaccat tattagaact ttcacgtaaa
     4321 catcaattta tgatttttga agatagaaaa tttgctgata ttggtaatac cgtgaagaaa
     4381 caatatattg gtggagttta taaaattagt agttgggcag atattactaa tgctcatggt
     4441 gtcactggga atggagtagt tgaaggatta aaacagggag ctaaagaaac caccaccaac
     4501 caagagccaa gagggttatt gatgttagct gaattatcat cagtgggatc attagcatat
     4561 ggagaatatt ctcaaaaaac tgttgaaatt gctaaatccg ataaggaatt tgttattgga
     4621 tttattgccc aacgtgatat gggtggacaa gaagaaggat ttgattggct tattatgaca
     4681 cctggagttg gattagatga taaaggtgat ggattaggac aacaatatag aactgttgat
     4741 gaagttgtta gcactggaac tgatattatc attgttggta gaggattgtt tggtaaagga
     4801 agagatccag atattgaagg taaaaggtat agagatgctg gttggaatgc ttatttgaaa
     4861 aagactggcc aattataatc agtactgaca ataaaaagat tcttgttttc aagaacttgt
     4921 catttgtata gtttttttat attgtagttg ttctatttta atcaaatgtt agcgtgattt
     4981 atattttttt tcgcctcgac atcatctgcc cagatgcgaa gttaagtgcg cagaaagtaa
     5041 tatcatgcgt caatcgtatg tgaatgctgg tcgctatact gctgtcgatt cgatactaac
     5101 gccgccatcc agtgtcgaaa acgagctcga attcatcgat gatatcagat ccactagtgg
     5161 cctatgctaa aaatttttgc agacattttt gatatatatt caattcatat tttatctctt
     5221 tttttttttt ttttttcaga cgtggaatgg tgggaaagcc aaaatgtcac tgaccgcggc
     5281 taaaagttaa aaaatacatg agacgacatt cttttatgta tatatattta cacatgtata
     5341 caaacaaata cacacgcaag tagagaggag ttggtggggg gaagatgcca ttatagagga
     5401 gaaaagaaca aagagagtga gcttgcaaga taaaatcact cgaaaaaaaa aatttcagtg
     5461 acttagccga taaatgaatg tgcttgcatt tttctaatta tccttcgtat cttctggctt
     5521 agtcacgggc caagcgtaag ggtgcttttc gggcataaca tacttgtgtt tttgcatata
     5581 ttccttcaat ccctttggac ctcttgatcc gtaggggtaa atttccggtg ttggaccgtc
     5641 cggacgctct atgtgcttca gtaatggggt gaatatgccc caactgatat ccaattcgtc
     5701 atctctgaca aagttggaat ggtcacccag tagggcgtct cttatcaaca cctcgtaagc
     5761 ctctggaatc caaaagtctt ggtacccagc ttttgttccc tttagtgagg gttaattgcg
     5821 cgcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
     5881 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
     5941 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     6001 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
     6061 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     6121 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     6181 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     6241 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     6301 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     6361 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     6421 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     6481 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     6541 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     6601 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     6661 aactacggct acactagaag gacagtattt ggtatctgcg ctctgctgaa gccagttacc
     6721 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     6781 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     6841 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     6901 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
     6961 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
     7021 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
     7081 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
     7141 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
     7201 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
     7261 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     7321 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     7381 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     7441 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     7501 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     7561 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
     7621 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
     7681 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
     7741 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
     7801 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
     7861 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac
     7921 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa
     7981 gtgccac
//
