LOCUS       sequence-128247-        6875 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Integration of GFP tag at Atg21 C terminus, use auxotrophic marker 
            URA3(Candida albicans).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6875)
  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 6875)
  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: LD121
FEATURES             Location/Qualifiers
     source          1..6875
                     /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             892..903
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     CDS             1206..1922
                     /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"
     terminator      1943..2130
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(1998..2022)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     promoter        2205..2548
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     terminator      3367..3564
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(4687..4707)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4687..4705)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4726..4742)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4726..4742)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4739..4761)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4750..4766
                     /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(4774..4780,4781..4798,4799..4804))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4819..4840
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4957..4974)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5128..5716)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5208..5227)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5887..6678,6679..6747))
                     /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     6510..6529
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6748..6852)
                     /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 ccgcggcttc aaggtcactg tttaaagaat ttagacgggg taccagattg tgcaatttgt
      721 atcagctagc tttcgataag agtatgacta tgattggatg tgtaggtgat acggacacta
      781 ttcatctttt caaacttgat gacgcctcta atagcctccc tggagataat tctagcaatg
      841 gccactggaa tgaagaggaa tatatactag catcaaattc aaatcccagt atgggaactc
      901 caaaggaaat acctctctcg aaaccaagaa ttgcgaatta cttctcgaaa aaaataaaat
      961 cttcaatacc aaaccagaac ctgagccgta actttgctta tataacagtt aatgaatcga
     1021 acaggagttg cttagggttt ccagacgaat ttcccaacca agtctatatt gcctctgatg
     1081 atggaacctt cagtatatat agtattccgt caaaacctgg tgaatgtgtg ctgactaaaa
     1141 ataataaatt tacagcttcg tacgctgcag gtcgacggat cggtgacggt gctggtttaa
     1201 ttaacatgtc taaaggtgaa gaattattca ctggtgttgt cccaattttg gttgaattag
     1261 atggtgatgt taatggtcac aaattttctg tctccggtga aggtgaaggt gatgctactt
     1321 acggtaaatt gaccttaaaa tttatttgta ctactggtaa attgccagtt ccatggccaa
     1381 ccttagtcac tactttaact tatggtgttc aatgtttttc tagataccca gatcatatga
     1441 aacaacatga ctttttcaag tctgccatgc cagaaggtta tgttcaagaa agaactattt
     1501 ttttcaaaga tgacggtaac tacaagacca gagctgaagt caagtttgaa ggtgatacct
     1561 tagttaatag aatcgaatta aaaggtattg attttaaaga agatggtaac attttaggtc
     1621 acaaattgga atacaactat aactctcaca atgtttacat catggctgac aaacaaaaga
     1681 atggtatcaa agttaacttc aaaattagac acaacattga agatggttct gttcaattag
     1741 ctgaccatta tcaacaaaat actccaattg gtgatggtcc agtcttgtta ccagacaacc
     1801 attacttatc cactcaatct gccttatcca aagatccaaa cgaaaagaga gaccacatgg
     1861 tcttgttaga atttgttact gctgctggta ttacccatgg tattgatgaa ttgtacaaat
     1921 aaggcgcgcc acttctaaat aagcgaattt cttatgattt atgattttta ttattaaata
     1981 agttataaaa aaaataagtg tatacaaatt ttaaagtgac tcttaggttt taaaacgaaa
     2041 attcttattc ttgagtaact ctttcctgta ggtcaggttg ctttctcagg tatagtatga
     2101 ggtcgctctt attgaccaca cctctaccgg cagatccgct agggataaca gggtaatata
     2161 gatctgttta gcttgcctcg tccccgccgg gtcacccggc cagcgacatg gaggcccaga
     2221 ataccctcct tgacagtctt gacgtgcgca gctcaggggc atgatgtgac tgtcgcccgt
     2281 acatttagcc catacatccc catgtataat catttgcatc catacatttt gatggccgca
     2341 cggcgcgaag caaaaattac ggctcctcgc tgcagacctg cgagcaggga aacgctcccc
     2401 tcacagacgc gttgaattgt ccccacgccg cgcccctgta gagaaatata aaaggttagg
     2461 atttgccact gaggttcttc tttcatatac ttccttttaa aatcttgcta ggatacagtt
     2521 ctcacatcac atccgaacat aaacaaccat gacagtcaac actaagacct atagtgagag
     2581 agcagaaact catgcctcac cagtagcaca gcgattattt cgattaatgg aactgaagaa
     2641 aaccaattta tgtgcatcaa ttgacgttga taccactaag gaattccttg aattaattga
     2701 taaattaggt ccttatgtat gcttaatcaa gactcatatt gatataatca atgatttttc
     2761 ctatgaatcc actattgaac cattattaga actttcacgt aaacatcaat ttatgatttt
     2821 tgaagataga aaatttgctg atattggtaa taccgtaaag aaacaatata ttggtggagt
     2881 ttataaaatt agtagttggg cagatattac caatgctcat ggtgtcactg ggaatggagt
     2941 ggttgaagga ttaaaacagg gagctaaaga aaccaccacc aaccaagagc caagagggtt
     3001 attgatgtta gctgaattat catcagtggg atcattagca tatggagaat attctcaaaa
     3061 aactgttgaa attgctaaat ccgataagga atttgttatt ggatttattg cccaacgtga
     3121 tatgggtggc caagaagaag gatttgattg gcttattatg acacctggag ttggattaga
     3181 tgataaaggt gatggattag gacaacaata tagaactgtt gatgaagttg ttagcactgg
     3241 aactgatatt atcattgttg gtagaggatt gtttggtaaa ggaagagatc cagatattga
     3301 aggtaaaagg tatagaaatg ctggttggaa tgcttatttg aaaaagactg gccaattata
     3361 atcagtactg acaataaaaa gattcttgtt ttcaagaact tgtcatttgt atagtttttt
     3421 tatattgtag ttgttctatt ttaatcaaat gttagcgtga tttatatttt ttttcgcctc
     3481 gacatcatct gcccagatgc gaagttaagt gcgcagaaag taatatcatg cgtcaatcgt
     3541 atgtgaatgc tggtcgctat actgctgtcg attcgatact aacgccgcca tccagtgtcg
     3601 aaaacgagct cgaattcatc gatgatatca gatccactag tggcctatgc taatgcatat
     3661 gtgcagtttt cataatctta atagatattg tacgtattca cgtgtgaatt gctaaagagg
     3721 gtagaaatct tcaaaacaag caatttgaaa atatttacat aggcaatggt acattctaga
     3781 aacacgaaaa gccaacaaga taactaattc tcaccaaaac catctcctat accaaggctt
     3841 catatgtgtt gattcattac tgccctttcc cctactagta ttgtgggttt tatcctgtac
     3901 aattaactca tttaattggt gttccaggat ttctacctct ttctttagct gactttctct
     3961 ctcctcatgc tctagtttcc accaagagaa gtgaagcagt aaataaacgg aagatcccat
     4021 tagaatgaac atccaaagag gcctaataaa tctttttaca gggatctcat gtcctgattc
     4081 ttcctcttgc gagaattttt tgagcatatc taactccttt ttgatattta attttgtagt
     4141 atgctcatta atcaaatcgc gtatcagttc agtatcaaca ccatccaaag aatccaaatt
     4201 agccaagtct tctaggctct tgatttcttt cttcggtgac gttgaaacag ctcttcgttg
     4261 ttggttgagg aggcgcctgt ggtaataggc aacggttgtt atgggaaaca atttagttct
     4321 actcaaagta atgagagcat tagagcgtct ttttaacata ttgtctatta tatgctttgg
     4381 ttcccatttc ctttctcaaa gtttacaaat ataacagagt gcttagcgaa tactttattt
     4441 tatgagtctt gccgttagtg accccaccca aaataagatg caaaccggta tacatattag
     4501 acatttccca cgtatacaaa tatattaagt aaataagcct cgtcgtgatg aacttaaata
     4561 catggtattg agagagttat aaacaggtag atagatatct taaataaaaa taagcagata
     4621 agtaacaatg tttaagaaaa cgatggggaa gcgctcgagg gggggcccgg tacccagctt
     4681 ttgttccctt tagtgagggt taattgcgcg cttggcgtaa tcatggtcat agctgtttcc
     4741 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg
     4801 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
     4861 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     4921 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     4981 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     5041 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     5101 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     5161 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     5221 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     5281 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     5341 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     5401 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     5461 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     5521 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg
     5581 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     5641 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     5701 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     5761 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     5821 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     5881 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     5941 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     6001 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     6061 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     6121 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     6181 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     6241 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     6301 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     6361 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     6421 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     6481 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     6541 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     6601 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     6661 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     6721 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     6781 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     6841 aggggttccg cgcacatttc cccgaaaagt gccac
//
