LOCUS       sequence-128293-        8034 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Integration of 2GFP tag at Atg18 C terminus, use auxotrophic marker 
            URA3(Candida albicans).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8034)
  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 8034)
  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: LD218
FEATURES             Location/Qualifiers
     source          1..8034
                     /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             1618..2331
                     /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             2368..3084
                     /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             2368..3081
                     /codon_start=1
                     /product="green fluorescent protein"
                     /label=GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     terminator      3105..3292
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(3160..3184)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     promoter        3367..3710
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     terminator      4529..4726
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(5846..5866)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5846..5864)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5885..5901)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5885..5901)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5898..5920)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5909..5925
                     /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(5933..5939,5940..5957,5958..5963))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5978..5999
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6116..6133)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6287..6875)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6367..6386)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7046..7837,7838..7906))
                     /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     7669..7688
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7907..8011)
                     /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 ccgcggtcga gctttactaa gagggatgat gcggatccaa caagcagtaa cggcggtaac
      721 agcagtataa taaagaatgg tgatgtgatc gtattcaact tggaaacatt acagccaacc
      781 atggtcatcg aagctcataa gggcgagatt gctgcaatgg caattagttt tgatgggaca
      841 ctaatggcta ccgcctctga taaaggtact atcatcaggg tctttgacat tgaaacgggt
      901 gataagatct accaattcag gagagggacg tacgcgacaa gaatttactc catatcattc
      961 agtgaagata gccagtactt ggcggttacc ggctcttcca aaaccgtgca tatcttcaaa
     1021 ttggggcatt caatgagcaa caataaacta gacagcgatg atagcaacat ggaagaagct
     1081 gcagccgatg attcatcgct cgataccacc agtatcgatg cgctgagtga cgaggaaaac
     1141 ccgacaagac tcgcaagaga accatatgtg gatgcatcaa gaaagacaat gggtaggatg
     1201 atacgttact cttctcaaaa gctatcccga agagctgcca gaacattggg tcagattttc
     1261 cccatcaaag ttacatcgtt gttggaatcc tcgcgccatt ttgcgtcttt gaaacttccc
     1321 gttgaaacca attcccatgt aatgaccata tcaagtatag gctctccaat agatatagac
     1381 acatccgagt atccggaact cttcgaaact ggcaattccg caagtacaga gtcctaccat
     1441 gagcctgtta tgaagatggt ccccatcagg gtcgtttcct cggatggata cctatacaac
     1501 tttgttatgg acccggagag aggcggcgat tgcttaatat tgtcacagta ttccatcttg
     1561 atggatgctt cgtacgctgc aggtcgacgg atcggtgacg gtgctggttt aattaacatg
     1621 tctaaaggtg aagaattatt cactggtgtt gtcccaattt tggttgaatt agatggtgat
     1681 gttaatggtc acaaattttc tgtctccggt gaaggtgaag gtgatgctac ttacggtaaa
     1741 ttgaccttaa aatttatttg tactactggt aaattgccag ttccatggcc aaccttagtc
     1801 actactttaa cttatggtgt tcaatgtttt tctagatacc cagatcatat gaaacaacat
     1861 gactttttca agtctgccat gccagaaggt tatgttcaag aaagaactat ttttttcaaa
     1921 gatgacggta actacaagac cagagctgaa gtcaagtttg aaggtgatac cttagttaat
     1981 agaatcgaat taaaaggtat tgattttaaa gaagatggta acattttagg tcacaaattg
     2041 gaatacaact ataactctca caatgtttac atcatggctg acaaacaaaa gaatggtatc
     2101 aaagttaact tcaaaattag acacaacatt gaagatggtt ctgttcaatt agctgaccat
     2161 tatcaacaaa atactccaat tggtgatggt ccagtcttgt taccagacaa ccattactta
     2221 tccactcaat ctgccttatc caaagatcca aacgaaaaga gagaccacat ggtcttgtta
     2281 gaatttgtta ctgctgctgg tattacccat ggtattgatg aattgtacaa aattccagtt
     2341 ccaacgcgtg atctagagaa gccattcatg tctaaaggtg aagaattatt cactggtgtt
     2401 gtcccaattt tggttgaatt agatggtgat gttaatggtc acaaattttc tgtctccggt
     2461 gaaggtgaag gtgatgctac ttacggtaaa ttgaccttaa aatttatttg tactactggt
     2521 aaattgccag ttccatggcc aaccttagtc actactttaa cttatggtgt tcaatgtttt
     2581 tctagatacc cagatcatat gaaacaacat gactttttca agtctgccat gccagaaggt
     2641 tatgttcaag aaagaactat ttttttcaaa gatgacggta actacaagac cagagctgaa
     2701 gtcaagtttg aaggtgatac cttagttaat agaatcgaat taaaaggtat tgattttaaa
     2761 gaagatggta acattttagg tcacaaattg gaatacaact ataactctca caatgtttac
     2821 atcatggctg acaaacaaaa gaatggtatc aaagttaact tcaaaattag acacaacatt
     2881 gaagatggtt ctgttcaatt agctgaccat tatcaacaaa atactccaat tggtgatggt
     2941 ccagtcttgt taccagacaa ccattactta tccactcaat ctgccttatc caaagatcca
     3001 aacgaaaaga gagaccacat ggtcttgtta gaatttgtta ctgctgctgg tattacccat
     3061 ggtatggatg aattatataa ataaggcgcg ccacttctaa ataagcgaat ttcttatgat
     3121 ttatgatttt tattattaaa taagttataa aaaaaataag tgtatacaaa ttttaaagtg
     3181 actcttaggt tttaaaacga aaattcttat tcttgagtaa ctctttcctg taggtcaggt
     3241 tgctttctca ggtatagtat gaggtcgctc ttattgacca cacctctacc ggcagatccg
     3301 ctagggataa cagggtaata tagatctgtt tagcttgcct cgtccccgcc gggtcacccg
     3361 gccagcgaca tggaggccca gaataccctc cttgacagtc ttgacgtgcg cagctcaggg
     3421 gcatgatgtg actgtcgccc gtacatttag cccatacatc cccatgtata atcatttgca
     3481 tccatacatt ttgatggccg cacggcgcga agcaaaaatt acggctcctc gctgcagacc
     3541 tgcgagcagg gaaacgctcc cctcacagac gcgttgaatt gtccccacgc cgcgcccctg
     3601 tagagaaata taaaaggtta ggatttgcca ctgaggttct tctttcatat acttcctttt
     3661 aaaatcttgc taggatacag ttctcacatc acatccgaac ataaacaacc atgacagtca
     3721 acactaagac ctatagtgag agagcagaaa ctcatgcctc accagtagca cagcgattat
     3781 ttcgattaat ggaactgaag aaaaccaatt tatgtgcatc aattgacgtt gataccacta
     3841 aggaattcct tgaattaatt gataaattag gtccttatgt atgcttaatc aagactcata
     3901 ttgatataat caatgatttt tcctatgaat ccactattga accattatta gaactttcac
     3961 gtaaacatca atttatgatt tttgaagata gaaaatttgc tgatattggt aataccgtaa
     4021 agaaacaata tattggtgga gtttataaaa ttagtagttg ggcagatatt accaatgctc
     4081 atggtgtcac tgggaatgga gtggttgaag gattaaaaca gggagctaaa gaaaccacca
     4141 ccaaccaaga gccaagaggg ttattgatgt tagctgaatt atcatcagtg ggatcattag
     4201 catatggaga atattctcaa aaaactgttg aaattgctaa atccgataag gaatttgtta
     4261 ttggatttat tgcccaacgt gatatgggtg gccaagaaga aggatttgat tggcttatta
     4321 tgacacctgg agttggatta gatgataaag gtgatggatt aggacaacaa tatagaactg
     4381 ttgatgaagt tgttagcact ggaactgata ttatcattgt tggtagagga ttgtttggta
     4441 aaggaagaga tccagatatt gaaggtaaaa ggtatagaaa tgctggttgg aatgcttatt
     4501 tgaaaaagac tggccaatta taatcagtac tgacaataaa aagattcttg ttttcaagaa
     4561 cttgtcattt gtatagtttt tttatattgt agttgttcta ttttaatcaa atgttagcgt
     4621 gatttatatt ttttttcgcc tcgacatcat ctgcccagat gcgaagttaa gtgcgcagaa
     4681 agtaatatca tgcgtcaatc gtatgtgaat gctggtcgct atactgctgt cgattcgata
     4741 ctaacgccgc catccagtgt cgaaaacgag ctcgaattca tcgatgatat cagatccact
     4801 agtggcctat gctgatcacg gaagtgtctc gcgctgcctt ccgtacgtca caacgcatac
     4861 acaatatagt ttatataaac tcctgcgtat aatctatcct tttgcataat attttcgtca
     4921 ccattatctt gttgcggtgt ttttatttaa aacaatagaa ctcgcctaaa ggggaaattt
     4981 tcgatataaa aattcaaaaa aatggctttc atggatcgag tatttgtttg tcgaaaaagg
     5041 atcactggag tgacgttact acgctacgaa gcctccttcc ggcttagccc tgttgattac
     5101 gaatttggat cagttggtat ttgattcctc ggaagagtta acttcacgcg ggtaaatcac
     5161 ttgtgatgcg gtatattctc tatacggcta atagatgaat cagggtgttt taaagtgcgt
     5221 ataaaccctt ttgctatttt cgtttatata attggcgttg ataaagagcc aatatctatt
     5281 gttgctacat agaggcagct ctcttagcaa aataaaaata caaaaagttc gacatgggct
     5341 tcagtagcgg taaatcaact aagaaaaagc ctctgctttt cgatatcaga cttaaaaatg
     5401 ttgacaacga tgtaatactc ctcaaaggtc ctccaaacga ggccccctcg gtgcttttat
     5461 ctggttgcat cgttttatcg attaacgaac ccatgcagat caaaagcata tcattgagac
     5521 tttatgggaa gatacaaata gacgtaccat tagagaggcc ccaggacgct agttcttcgt
     5581 cgttgtcttc atcgccgcca aagatcagaa agtacaacaa agttttttat aattacgcat
     5641 gggataatgt taacctcaag gagtatctga gtggtttaag agggcaatct ggccttgcgg
     5701 gcagtagctc atcaagtaat atcttgggca ctcgccaaag agctcagtcc acaagttcct
     5761 tgaagtcttt aaaggggtcc tcctcaccct cttcatgtac tttagataag ggctcgaggg
     5821 ggggcccggt acccagcttt tgttcccttt agtgagggtt aattgcgcgc ttggcgtaat
     5881 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac
     5941 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
     6001 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     6061 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc
     6121 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     6181 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     6241 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     6301 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     6361 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     6421 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     6481 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     6541 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     6601 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     6661 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     6721 ctagaaggac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     6781 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     6841 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     6901 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     6961 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     7021 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     7081 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     7141 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     7201 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     7261 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     7321 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     7381 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     7441 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     7501 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     7561 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     7621 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     7681 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     7741 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     7801 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     7861 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     7921 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     7981 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccac
//
