LOCUS       sequence-121959-        7134 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Integration of GFP tag at Atg20 C terminus, use auxotrophic marker 
            URA3(Candida albicans).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7134)
  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 7134)
  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: LD120
FEATURES             Location/Qualifiers
     source          1..7134
                     /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             1938..2654
                     /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             1938..2651
                     /codon_start=1
                     /product="green fluorescent protein"
                     /label=GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     terminator      2675..2862
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2730..2754)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     promoter        2937..3280
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     terminator      4099..4296
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(4946..4966)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4946..4964)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4985..5001)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4985..5001)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4998..5020)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5009..5025
                     /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(5033..5039,5040..5057,5058..5063))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5078..5099
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5216..5233)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5387..5975)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5467..5486)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6146..6937,6938..7006))
                     /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     6769..6788
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7007..7111)
                     /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 ccgcggagta tttgctgccc tcagaaggtt ctggttcagt agatctatca ttgtctgtga
      721 tacatgcgtc ggtcaataat agtgatgaaa agttaatacg gcataggata agaatgctca
      781 ctgagttctt gaacaaactt ttgacgaacg aagaaatcac aaaaacatct ataattaccg
      841 attttttgga tcctaataac cataattggc atgaatttgt taacagttcg tccacttttt
      901 catctttacc gaagagcatc ctccagtgta atccactaga tcccactaac accactagaa
      961 ttcatgccat gttaccaata ccgggctcct cgtcacagtt attgttgaat aaggagtcca
     1021 acgacaaaaa aatggataag gaaagaagta agtcgtttac taatattgaa caggattaca
     1081 agcaatatga gaacttgtta gataacggga tttataagta taatagacga accacgaaga
     1141 cttatcacga tttgaaatcg gattacaatg aaatcggtga ggtatttgca cagtttgccc
     1201 atgaacaggc tcaagttggg gaattggccg aacagctttc ttacttaagc aatgcctttt
     1261 cggggtcctc gatttcttta gaaaaattag ttggaagact atactacaat ataaatgagc
     1321 ctttaaatga atctgttcat atggcaactt ctgcaagaga attgatcaaa tacagaaaat
     1381 tgaaatacct gcaaaatgaa atgattaaaa agtctctgaa ttccaaaaga gcacagttgg
     1441 agaagctgga agcgcaaaac aatgaatata aggatgttga caaaataatt gacaacgaaa
     1501 tgtctaaaag tcatactatt aatctggaac gccctaataa taatacaggt agcggtggga
     1561 aatcatatgg tggcaaacta ttcaatggat ttaacaaatt ggcatcaatg gtaaaagatt
     1621 ctgtgaagta tcaagaaaca gatccgcata ccgcaagtat caatttgaag aaagaaattg
     1681 aacaactgtc tgaatcattg gaagttactg agaatgattt ggaggtcatt tcaaaagtca
     1741 tcaaaaatga ccagctacct aaattttcta aagaaaggga agtagattta tcagaaattt
     1801 taaagcatta ttccagatat atgagaaatt atgcccgtca aaatttagaa atttggaaag
     1861 aagtcaaaag acatcaagat tttgcagctt cgtacgctgc aggtcgacgg atcggtgacg
     1921 gtgctggttt aattaacatg tctaaaggtg aagaattatt cactggtgtt gtcccaattt
     1981 tggttgaatt agatggtgat gttaatggtc acaaattttc tgtctccggt gaaggtgaag
     2041 gtgatgctac ttacggtaaa ttgaccttaa aatttatttg tactactggt aaattgccag
     2101 ttccatggcc aaccttagtc actactttaa cttatggtgt tcaatgtttt tctagatacc
     2161 cagatcatat gaaacaacat gactttttca agtctgccat gccagaaggt tatgttcaag
     2221 aaagaactat ttttttcaaa gatgacggta actacaagac cagagctgaa gtcaagtttg
     2281 aaggtgatac cttagttaat agaatcgaat taaaaggtat tgattttaaa gaagatggta
     2341 acattttagg tcacaaattg gaatacaact ataactctca caatgtttac atcatggctg
     2401 acaaacaaaa gaatggtatc aaagttaact tcaaaattag acacaacatt gaagatggtt
     2461 ctgttcaatt agctgaccat tatcaacaaa atactccaat tggtgatggt ccagtcttgt
     2521 taccagacaa ccattactta tccactcaat ctgccttatc caaagatcca aacgaaaaga
     2581 gagaccacat ggtcttgtta gaatttgtta ctgctgctgg tattacccat ggtatggatg
     2641 aattgtacaa ataaggcgcg ccacttctaa ataagcgaat ttcttatgat ttatgatttt
     2701 tattattaaa taagttataa aaaaaataag tgtatacaaa ttttaaagtg actcttaggt
     2761 tttaaaacga aaattcttat tcttgagtaa ctctttcctg taggtcaggt tgctttctca
     2821 ggtatagtat gaggtcgctc ttattgacca cacctctacc ggcagatccg ctagggataa
     2881 cagggtaata tagatctgtt tagcttgcct cgtccccgcc gggtcacccg gccagcgaca
     2941 tggaggccca gaataccctc cttgacagtc ttgacgtgcg cagctcaggg gcatgatgtg
     3001 actgtcgccc gtacatttag cccatacatc cccatgtata atcatttgca tccatacatt
     3061 ttgatggccg cacggcgcga agcaaaaatt acggctcctc gctgcagacc tgcgagcagg
     3121 gaaacgctcc cctcacagac gcgttgaatt gtccccacgc cgcgcccctg tagagaaata
     3181 taaaaggtta ggatttgcca ctgaggttct tctttcatat acttcctttt aaaatcttgc
     3241 taggatacag ttctcacatc acatccgaac ataaacaacc atgacagtca acactaagac
     3301 ctatagtgag agagcagaaa ctcatgcctc accagtagca cagcgattat ttcgattaat
     3361 ggaactgaag aaaaccaatt tatgtgcatc aattgacgtt gataccacta aggaattcct
     3421 tgaattaatt gataaattag gtccttatgt atgcttaatc aagactcata ttgatataat
     3481 caatgatttt tcctatgaat ccactattga accattatta gaactttcac gtaaacatca
     3541 atttatgatt tttgaagata gaaaatttgc tgatattggt aataccgtaa agaaacaata
     3601 tattggtgga gtttataaaa ttagtagttg ggcagatatt accaatgctc atggtgtcac
     3661 tgggaatgga gtggttgaag gattaaaaca gggagctaaa gaaaccacca ccaaccaaga
     3721 gccaagaggg ttattgatgt tagctgaatt atcatcagtg ggatcattag catatggaga
     3781 atattctcaa aaaactgttg aaattgctaa atccgataag gaatttgtta ttggatttat
     3841 tgcccaacgt gatatgggtg gccaagaaga aggatttgat tggcttatta tgacacctgg
     3901 agttggatta gatgataaag gtgatggatt aggacaacaa tatagaactg ttgatgaagt
     3961 tgttagcact ggaactgata ttatcattgt tggtagagga ttgtttggta aaggaagaga
     4021 tccagatatt gaaggtaaaa ggtatagaaa tgctggttgg aatgcttatt tgaaaaagac
     4081 tggccaatta taatcagtac tgacaataaa aagattcttg ttttcaagaa cttgtcattt
     4141 gtatagtttt tttatattgt agttgttcta ttttaatcaa atgttagcgt gatttatatt
     4201 ttttttcgcc tcgacatcat ctgcccagat gcgaagttaa gtgcgcagaa agtaatatca
     4261 tgcgtcaatc gtatgtgaat gctggtcgct atactgctgt cgattcgata ctaacgccgc
     4321 catccagtgt cgaaaacgag ctcgaattca tcgatgatat cagatccact agtggcctat
     4381 gctagtatta tgagcatttt tttttagcgt tatagcatac ttgatatata tatatataaa
     4441 caacgtgaag atcaaataac gtcaaatgta ttaaatagac taaaatatta tgcacttctc
     4501 ttcggattta ttgctcgcgg gctggttgac gttacatggt ccattccact gaaatgtttt
     4561 aagagatgcc tatatggcca actcaaagtt ttgaaaaagg gaactaggtt cacataatat
     4621 ggggcattta gtgtttgcag atgattatgc tccatcgcag agattatcgc tagtgccagt
     4681 tgggaaggta tgatgtcagg agccaataac ttagatggtg tagggacatc aataaacata
     4741 ttagttttga tttttcccgg gcacaccagt agtgttttta ttccagtttt attacattcc
     4801 gtggacaaac ttctgaaatg cctgctcata cacttgtgaa aaccaatcat tgctcctttt
     4861 gatgcaccat acgatgtcag ccttgcagga gtcagttctc cgaggactga agctcgaggg
     4921 ggggcccggt acccagcttt tgttcccttt agtgagggtt aattgcgcgc ttggcgtaat
     4981 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac
     5041 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
     5101 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     5161 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc
     5221 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     5281 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     5341 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     5401 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     5461 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     5521 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     5581 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     5641 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     5701 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     5761 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     5821 ctagaaggac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     5881 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     5941 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     6001 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     6061 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     6121 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     6181 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     6241 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     6301 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     6361 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     6421 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     6481 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     6541 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     6601 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     6661 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     6721 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     6781 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     6841 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     6901 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     6961 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     7021 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     7081 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccac
//
