LOCUS       sequence-128279-        7268 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Integration of GFP tag at Atg27 C terminus, use auxotrophic marker 
            URA3(Candida albicans).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7268)
  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 7268)
  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: LD127
FEATURES             Location/Qualifiers
     source          1..7268
                     /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             1531..2247
                     /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      2268..2455
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2323..2347)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     protein_bind    2738..2771
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     terminator      3726..3923
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     protein_bind    3977..4010
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(5080..5100)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5080..5098)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5119..5135)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5119..5135)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5132..5154)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5143..5159
                     /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(5167..5173,5174..5191,5192..5197))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5212..5233
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5350..5367)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5521..6109)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5601..5620)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6280..7071,7072..7140))
                     /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     6903..6922
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7141..7245)
                     /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 ccgcggatgg tatcgaagac ttggatatgt ggcttcatca gtataattac agtggtacag
      721 gccttgtcct gcgagaagca tgatgtattg aaaaagtatc aggtgggaaa atttagctca
      781 ctaacttcta cggaaaggga tactccgcca agcacaacta ttgaaaagtg gtggataaac
      841 gtttgcgaag agcataacgt agaacctcct gaagaatgta aaaaaaatga catgctatgt
      901 ggtttaacag atgtcatctt gcccggtaag gatgctatca ccactcaaat tatagatttt
      961 gacaaaaaca ttggcttcaa tgtcgaggaa actgagagtg cgcttacatt gacactaaaa
     1021 ggcgctacgt ggggcgccaa ttcttttgac gcaaaactag aatttcagtg taatgacaat
     1081 atgaaacaag acgagctgac tagtcacaca tgggccgata aaagtatcca attgacccta
     1141 aaagggccat ctggctgttt gaagagcaaa gatgatgaca aaaaaaatgg cgacggcgat
     1201 aatggtaagg atggtgacag cgaggggaaa aagccagcta agaaggcagg cgggacttcg
     1261 tggttcactt ggttattcct atacgctcta ctgtttacat tgatatacct gatggttgta
     1321 tcattcttga ataccagagg aggctctttt caagatttcc gtgctgagtt cattcaacgt
     1381 tctactcaat ttctcacttc cttacctgaa ttctgcaagg aagtggtttc gagaatctta
     1441 ggcagaagta ccgcccaaag aggcggttat agcgccgttg cttcgtacgc tgcaggtcga
     1501 cggatcggtg acggtgctgg tttaattaac atgtctaaag gtgaagaatt attcactggt
     1561 gttgtcccaa ttttggttga attagatggt gatgttaatg gtcacaaatt ttctgtctcc
     1621 ggtgaaggtg aaggtgatgc tacttacggt aaattgacct taaaatttat ttgtactact
     1681 ggtaaattgc cagttccatg gccaacctta gtcactactt taacttatgg tgttcaatgt
     1741 ttttctagat acccagatca tatgaaacaa catgactttt tcaagtctgc catgccagaa
     1801 ggttatgttc aagaaagaac tatttttttc aaagatgacg gtaactacaa gaccagagct
     1861 gaagtcaagt ttgaaggtga taccttagtt aatagaatcg aattaaaagg tattgatttt
     1921 aaagaagatg gtaacatttt aggtcacaaa ttggaataca actataactc tcacaatgtt
     1981 tacatcatgg ctgacaaaca aaagaatggt atcaaagtta acttcaaaat tagacacaac
     2041 attgaagatg gttctgttca attagctgac cattatcaac aaaatactcc aattggtgat
     2101 ggtccagtct tgttaccaga caaccattac ttatccactc aatctgcctt atccaaagat
     2161 ccaaacgaaa agagagacca catggtcttg ttagaatttg ttactgctgc tggtattacc
     2221 catggtattg atgaattgta caaataaggc gcgccacttc taaataagcg aatttcttat
     2281 gatttatgat ttttattatt aaataagtta taaaaaaaat aagtgtatac aaattttaaa
     2341 gtgactctta ggttttaaaa cgaaaattct tattcttgag taactctttc ctgtaggtca
     2401 ggttgctttc tcaggtatag tatgaggtcg ctcttattga ccacacctct accggcagat
     2461 ccgctaggga taacagggta atatagatct gtttagcttg cctcgtcccc gccgggtcac
     2521 ccggccagcg acatggaggc ccagaatacc ctccttgaca gtcttgacgt gcgcagctca
     2581 ggggcatgat gtgactgtcg cccgtacatt tagcccatac atccccatgt ataatcattt
     2641 gcatccatac attttgatgg ccgcacggcg cgaagcaaaa attacggctc ctcgctgcag
     2701 acctgcgagc agggaaacgc tcccctcaca gacgcgtata acttcgtata atgtatgcta
     2761 tacgaagtta ttgaattgtc cccacgccgc gcccctgtag agaaatataa aaggttagga
     2821 tttgccactg aggttcttct ttcatatact tccttttaaa atcttgctag gatacagttc
     2881 tcacatcaca tccgaacata aacaaccatg acagtcaaca ctaagaccta tagtgagaga
     2941 gcagaaactc atgcctcacc agtagcacaa cgattatttc gattaatgga actgaagaaa
     3001 accaatttat gtgcatcaat tgatgttgat accactaagg aattccttga attaattgat
     3061 aaattgggtc cttatgtatg cttaatcaag actcatattg atataatcaa tgatttttcc
     3121 tatgaatcca ctattgaacc attattagaa ctttcacgta aacatcaatt tatgattttt
     3181 gaagatagaa aatttgctga tattggtaat accgtgaaga aacaatatat tggtggagtt
     3241 tataaaatta gtagttgggc agatattact aatgctcatg gtgtcactgg gaatggagta
     3301 gttgaaggat taaaacaggg agctaaagaa accaccacca accaagagcc aagagggtta
     3361 ttgatgttag ctgaattatc atcagtggga tcattagcat atggagaata ttctcaaaaa
     3421 actgttgaaa ttgctaaatc cgataaggaa tttgttattg gatttattgc ccaacgtgat
     3481 atgggtggac aagaagaagg atttgattgg cttattatga cacctggagt tggattagat
     3541 gataaaggtg atggattagg acaacaatat agaactgttg atgaagttgt tagcactgga
     3601 actgatatta tcattgttgg tagaggattg tttggtaaag gaagagatcc agatattgaa
     3661 ggtaaaaggt atagagatgc tggttggaat gcttatttga aaaagactgg ccaattataa
     3721 tcagtactga caataaaaag attcttgttt tcaagaactt gtcatttgta tagttttttt
     3781 atattgtagt tgttctattt taatcaaatg ttagcgtgat ttatattttt tttcgcctcg
     3841 acatcatctg cccagatgcg aagttaagtg cgcagaaagt aatatcatgc gtcaatcgta
     3901 tgtgaatgct ggtcgctata ctgctgtcga ttcgatacta acgccgccat ccagtgtcga
     3961 aaacgagctc gaattcataa cttcgtataa tgtatgctat acgaagttat atcgatgata
     4021 tcagatccac tagtggccta tgctaaataa gtttactggc ttacaattcg atatttttgc
     4081 aacagcagtg ctaacctttt attatattta tataaactct aagcttttat atctacatta
     4141 tactaataaa aaaaacttgt tcccgtagta agttttccgg ctaagaaagg aaaggctatt
     4201 gccgctttct tttaattctt catcaatgcc tttagattgt ctattgtttt ttcaacagta
     4261 gtttctaaat agttcttctt tatctttaat gtttttcttt gatccagaag aacagtttcg
     4321 tcatgtgata aatcattaac gtatttggat ttatcctgta agataaacga tttaccgcaa
     4381 gatctccata ctttgtccgt tggataagac tctagttctt tctttgtcaa ttcagcaagc
     4441 ttttcttgtc tgtccaaata tgctagctgc tgattgacca tatccaattg tgtcctggcg
     4501 tttcttaagc tcactgtcat ttcttgtgct atctgtgaca tgtttgtaag tatgtattat
     4561 tgattcaatt tagtaaagag ctctgtagat agttctttaa ttcccatctg ctcatctgag
     4621 aaatttcttc actgttactt tttaattgct atatctttaa actcattcat tttcttcaga
     4681 gaaaagaaaa aaattaaggt aaaggaaaag gctaaataag aacaaaacac ttgaaggcaa
     4741 acgaaaccct acaaaagaga atacggaatg ctgccatcat taaggaaggg atgcttcatt
     4801 gtgaattcca taagattgaa actgccccga ttctactcat taaatgccca gccactgggg
     4861 acagacaaca cgattgagaa taacactcct acggaaacaa atagattgag caaaacttca
     4921 cagaagtttt gggaaaaggt gtcactaaat agggatgttg agaaaggaaa gattgctcta
     4981 caattagatg gcaggactat aaaaactcct ctaggaaatg gaattatagt tgataatgca
     5041 aagtctctcg agggggggcc cggtacccag cttttgttcc ctttagtgag ggttaattgc
     5101 gcgcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat
     5161 tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag
     5221 ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
     5281 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
     5341 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     5401 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     5461 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     5521 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     5581 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     5641 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     5701 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     5761 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     5821 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     5881 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     5941 taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac
     6001 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     6061 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     6121 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     6181 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     6241 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     6301 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     6361 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     6421 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     6481 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     6541 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     6601 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     6661 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     6721 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     6781 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     6841 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     6901 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     6961 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     7021 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     7081 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     7141 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     7201 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     7261 agtgccac
//
