LOCUS       sequence-128297-        7434 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Integration of 2GFP tag at Atg23 C terminus, use auxotrophic marker 
            URA3(Candida albicans).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7434)
  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 7434)
  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: LD223
FEATURES             Location/Qualifiers
     source          1..7434
                     /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"
     CDS             1277..1990
                     /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             2027..2743
                     /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             2027..2740
                     /codon_start=1
                     /product="green fluorescent protein"
                     /label=GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     terminator      2764..2951
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2819..2843)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     promoter        3026..3369
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     terminator      4188..4385
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(5246..5266)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5246..5264)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5285..5301)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5285..5301)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5298..5320)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5309..5325
                     /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(5333..5339,5340..5357,5358..5363))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5378..5399
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5516..5533)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5687..6275)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5767..5786)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6446..7237,7238..7306))
                     /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     7069..7088
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7307..7411)
                     /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 ccgcggaact atcaacatga aaaattttgt tcatatgaag gatctcatag aactgaaaat
      721 tgaagatcta caagagcaac taatgagaaa taaaaatgag tcttctacag tactaacgca
      781 acgtgaatta tggctagatt gtcaaaaaaa agtaggagat ttggaaagca aattaataac
      841 caaacttcgg agcagcagca actccaaaat acctccaaat gaaatgtcag aaatgatcaa
      901 ttctacaata cagtacctaa ataaccttct tgactcttct gatgaaaagc tcacgacaac
      961 tttaatatca aacgaaaggg acgtactttc gaaagcttgt gaagaactcc atagcgaaag
     1021 taccactgca caagatgggt caagcgcact accttctaaa ccaattgata tacacaaaag
     1081 ccataaaggg tcgaacgcga gttcaaattt aaaacagccc tcgactccct cattcttggt
     1141 tgctagcaaa tcgccaccaa agattggcat ttctgaaagt gtagtaaacg ctaataaaaa
     1201 tgatgccata tcaaaaaaag ttgaagcttc gtacgctgca ggtcgacgga tcggtgacgg
     1261 tgctggttta attaacatgt ctaaaggtga agaattattc actggtgttg tcccaatttt
     1321 ggttgaatta gatggtgatg ttaatggtca caaattttct gtctccggtg aaggtgaagg
     1381 tgatgctact tacggtaaat tgaccttaaa atttatttgt actactggta aattgccagt
     1441 tccatggcca accttagtca ctactttaac ttatggtgtt caatgttttt ctagataccc
     1501 agatcatatg aaacaacatg actttttcaa gtctgccatg ccagaaggtt atgttcaaga
     1561 aagaactatt tttttcaaag atgacggtaa ctacaagacc agagctgaag tcaagtttga
     1621 aggtgatacc ttagttaata gaatcgaatt aaaaggtatt gattttaaag aagatggtaa
     1681 cattttaggt cacaaattgg aatacaacta taactctcac aatgtttaca tcatggctga
     1741 caaacaaaag aatggtatca aagttaactt caaaattaga cacaacattg aagatggttc
     1801 tgttcaatta gctgaccatt atcaacaaaa tactccaatt ggtgatggtc cagtcttgtt
     1861 accagacaac cattacttat ccactcaatc tgccttatcc aaagatccaa acgaaaagag
     1921 agaccacatg gtcttgttag aatttgttac tgctgctggt attacccatg gtattgatga
     1981 attgtacaaa attccagttc caacgcgtga tctagagaag ccattcatgt ctaaaggtga
     2041 agaattattc actggtgttg tcccaatttt ggttgaatta gatggtgatg ttaatggtca
     2101 caaattttct gtctccggtg aaggtgaagg tgatgctact tacggtaaat tgaccttaaa
     2161 atttatttgt actactggta aattgccagt tccatggcca accttagtca ctactttaac
     2221 ttatggtgtt caatgttttt ctagataccc agatcatatg aaacaacatg actttttcaa
     2281 gtctgccatg ccagaaggtt atgttcaaga aagaactatt tttttcaaag atgacggtaa
     2341 ctacaagacc agagctgaag tcaagtttga aggtgatacc ttagttaata gaatcgaatt
     2401 aaaaggtatt gattttaaag aagatggtaa cattttaggt cacaaattgg aatacaacta
     2461 taactctcac aatgtttaca tcatggctga caaacaaaag aatggtatca aagttaactt
     2521 caaaattaga cacaacattg aagatggttc tgttcaatta gctgaccatt atcaacaaaa
     2581 tactccaatt ggtgatggtc cagtcttgtt accagacaac cattacttat ccactcaatc
     2641 tgccttatcc aaagatccaa acgaaaagag agaccacatg gtcttgttag aatttgttac
     2701 tgctgctggt attacccatg gtatggatga attatataaa taaggcgcgc cacttctaaa
     2761 taagcgaatt tcttatgatt tatgattttt attattaaat aagttataaa aaaaataagt
     2821 gtatacaaat tttaaagtga ctcttaggtt ttaaaacgaa aattcttatt cttgagtaac
     2881 tctttcctgt aggtcaggtt gctttctcag gtatagtatg aggtcgctct tattgaccac
     2941 acctctaccg gcagatccgc tagggataac agggtaatat agatctgttt agcttgcctc
     3001 gtccccgccg ggtcacccgg ccagcgacat ggaggcccag aataccctcc ttgacagtct
     3061 tgacgtgcgc agctcagggg catgatgtga ctgtcgcccg tacatttagc ccatacatcc
     3121 ccatgtataa tcatttgcat ccatacattt tgatggccgc acggcgcgaa gcaaaaatta
     3181 cggctcctcg ctgcagacct gcgagcaggg aaacgctccc ctcacagacg cgttgaattg
     3241 tccccacgcc gcgcccctgt agagaaatat aaaaggttag gatttgccac tgaggttctt
     3301 ctttcatata cttcctttta aaatcttgct aggatacagt tctcacatca catccgaaca
     3361 taaacaacca tgacagtcaa cactaagacc tatagtgaga gagcagaaac tcatgcctca
     3421 ccagtagcac agcgattatt tcgattaatg gaactgaaga aaaccaattt atgtgcatca
     3481 attgacgttg ataccactaa ggaattcctt gaattaattg ataaattagg tccttatgta
     3541 tgcttaatca agactcatat tgatataatc aatgattttt cctatgaatc cactattgaa
     3601 ccattattag aactttcacg taaacatcaa tttatgattt ttgaagatag aaaatttgct
     3661 gatattggta ataccgtaaa gaaacaatat attggtggag tttataaaat tagtagttgg
     3721 gcagatatta ccaatgctca tggtgtcact gggaatggag tggttgaagg attaaaacag
     3781 ggagctaaag aaaccaccac caaccaagag ccaagagggt tattgatgtt agctgaatta
     3841 tcatcagtgg gatcattagc atatggagaa tattctcaaa aaactgttga aattgctaaa
     3901 tccgataagg aatttgttat tggatttatt gcccaacgtg atatgggtgg ccaagaagaa
     3961 ggatttgatt ggcttattat gacacctgga gttggattag atgataaagg tgatggatta
     4021 ggacaacaat atagaactgt tgatgaagtt gttagcactg gaactgatat tatcattgtt
     4081 ggtagaggat tgtttggtaa aggaagagat ccagatattg aaggtaaaag gtatagaaat
     4141 gctggttgga atgcttattt gaaaaagact ggccaattat aatcagtact gacaataaaa
     4201 agattcttgt tttcaagaac ttgtcatttg tatagttttt ttatattgta gttgttctat
     4261 tttaatcaaa tgttagcgtg atttatattt tttttcgcct cgacatcatc tgcccagatg
     4321 cgaagttaag tgcgcagaaa gtaatatcat gcgtcaatcg tatgtgaatg ctggtcgcta
     4381 tactgctgtc gattcgatac taacgccgcc atccagtgtc gaaaacgagc tcgaattcat
     4441 cgatgatatc agatccacta gtggcctatg ctgaaatgca aatagctaga gtagccatga
     4501 ggataatgta aatttaaaca ttgtatgccc acgtaatagt caaactcttt cacgtcaaat
     4561 tgaaaaaaaa tgttcttttt gctgaaataa tgaaaaataa tgaaaaataa tgttgctatt
     4621 aaattcaata taaatgtata caccaatata tatgcaggta taattcctaa cacttttact
     4681 tcaagatcat gatctaggct ttcttttttt tgggataaat gggctagatg aagcattccg
     4741 tctcgaactg ttgcgtgtgc ctgtgggccc actaccaggt cctttgaatg ctgaagattg
     4801 tggttgcgga tgggggttta aatcatatgg atcttctgcc tcagcaggat agctgtcttg
     4861 cggatgggcg ggcagagttt catgcagttc gcctgtaccc acaggagccg ttgagtattg
     4921 attcgcatta cccacatgtt gagagagggt cctatttgct ccattctgtc cattttgatt
     4981 gggttcaccg taatcggaag cagtcaggat cctcgaagaa ccaccggata aaacactatt
     5041 tgaacttctg tgttttgtaa tagcagcagg tacctgcgtc tgctttgctt gcgaaacttg
     5101 tggatcgtcg tatatactag cgctgaaatt ttggttatga gaagccaccc tgttgttacg
     5161 accatttgct tcacttttca ttaatgaaga atcggagatt gacggcgtat aactcgaggg
     5221 ggggcccggt acccagcttt tgttcccttt agtgagggtt aattgcgcgc ttggcgtaat
     5281 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac
     5341 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
     5401 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     5461 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc
     5521 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     5581 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     5641 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     5701 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     5761 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     5821 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     5881 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     5941 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     6001 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     6061 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     6121 ctagaaggac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     6181 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     6241 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     6301 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     6361 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     6421 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     6481 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     6541 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     6601 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     6661 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     6721 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     6781 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     6841 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     6901 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     6961 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     7021 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     7081 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     7141 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     7201 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     7261 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     7321 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     7381 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccac
//
