LOCUS       sequence-240353-        7246 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expressing S. pombe GFP-Atg17.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7246)
  AUTHORS   Nanji T, Liu X, Chew LH, Li FK, Biswas M, Yu ZQ, Lu S, Dong MQ, Du 
            LL, Klionsky DJ, Yip CK
  TITLE     Conserved and unique features of the fission yeast core Atg1 
            complex.
  JOURNAL   Autophagy. 2017;13(12):2018-2027. doi: 
            10.1080/15548627.2017.1382782. Epub 2017 Nov 23.
  PUBMED    28976798
REFERENCE   2  (bases 1 to 7246)
  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: "2017"; volume: "13(12):2018-2027. doi"; pages: 
            " 10.1080/15548627.2017.1382782. Epub 2017 Nov 23"
COMMENT     Sequence Label: pCuGFP-SpAtg17(416)
FEATURES             Location/Qualifiers
     source          1..7246
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    105..126
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(141..146,147..164,165..171)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    179..195
                     /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)."
     primer_bind     184..206
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     203..219
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     203..219
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     238..258
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        240..258
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        277..511
                     /gene="S. cerevisiae CUP1"
                     /label=CUP1 promoter
                     /note="inducible metallothionein promoter, regulated by 
                     Cu2+"
     CDS             519..1232
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(549..577)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     1172..1192
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      2481..2728
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(2497..2515)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(2746..2765)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2747..2765)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2775..2792)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2775..2791)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2784..2806)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2932..3387
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3019..3038)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3229..3250
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3518..4321)
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     promoter        complement(4322..4537)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(4561..4580)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4680..4702
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4740..4758)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    4799..5302
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        5339..5443
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5444..5512,5513..6304)
                     /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     complement(5662..5681)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6475..7063
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6964..6983
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7217..7234
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 cgcccaatac gcaaaccgcc tctccccgcg cgttggccga ttcattaatg cagctggcac
       61 gacaggtttc ccgactggaa agcgggcagt gagcgcaacg caattaatgt gagttagctc
      121 actcattagg caccccaggc tttacacttt atgcttccgg ctcgtatgtt gtgtggaatt
      181 gtgagcggat aacaatttca cacaggaaac agctatgacc atgattacgc caagcgcgca
      241 attaaccctc actaaaggga acaaaagctg gagctcctag ttagaaaaag acatttttgc
      301 tgtcagtcac tgtcaagaga ttcttttgct ggcatttctt ctagaagcaa aaagagcgat
      361 gcgtcttttc cgctgaaccg ttccagcaaa aaagactacc aacgcaatat ggattgtcag
      421 aatcatataa aagagaagca aataactcct tgtcttgtat caattgcatt ataatatctt
      481 cttgttagtg caatatcata tagaagtcat cgactagtat gagtaaagga gaagaacttt
      541 tcactggagt tgtcccaatt cttgttgaat tagatggtga tgttaatggg cacaaatttt
      601 ctgtcagtgg agagggtgaa ggtgatgcaa catacggaaa acttaccctt aaatttattt
      661 gcactactgg aaaactacct gttccatggc caacacttgt cactactttc acttatggtg
      721 ttcaatgctt ttcaagatac ccagatcata tgaaacggca tgactttttc aagagtgcca
      781 tgcccgaagg ttatgtacag gaaagaacta tatttttcaa agatgacggg aactacaaga
      841 cacgtgctga agtcaagttt gaaggtgata cccttgttaa tagaatcgag ttaaaaggta
      901 ttgattttaa agaagatgga aacattcttg gacacaaatt ggaatacaac tataactcac
      961 acaatgtata catcatggca gacaaacaaa agaatggaat caaagttaac ttcaaaatta
     1021 gacacaacat tgaagatgga agcgttcaac tagcagacca ttatcaacaa aatactccaa
     1081 ttggcgatgg ccctgtcctt ttaccagaca accattacct gtccacacaa tctgcccttt
     1141 cgaaagatcc caacgaaaag agagaccaca tggtccttct tgagtttgta acagctgctg
     1201 ggattacaca tggcatggat gaactataca aacccgggat ggaattgttg cagcagtgga
     1261 ctcagcaagc caaagcagct cttactcaag ccaggcaact gtgtggtgat gcacataagt
     1321 ttaatgaaga tgcaaaaact gatttgagaa attccataaa gcagcatcaa cagctaaaag
     1381 agttggcgaa gctcacagca tctcaatgta ctcgtttaga ctcttcaaca gctcttatta
     1441 agcagttatt agatctagtt caaaattatc caacatttaa tcaattaaac gtccttcatg
     1501 atcgattgga atcgtcactt aaacgtcttc gtgactgtac attagatcca gccttaggaa
     1561 gcgaatatac caatctttat gcatttgtag atgatactgc attagaagat ttgaaaacca
     1621 ggctccgagg cgtaactgat ggtgtttgga atgctttcga aaagctagcg ggtttactgg
     1681 aagaagattt gtgtgccaat tatcataaac gtttagaagc tgtttcatta gacttcttac
     1741 ctcccgctta taacgatact gccgaagaac ttgcggattt gcttttgcaa gttgctcaac
     1801 attacgatca atgctccgaa gctctcaata tttatgacac cctttctgat gcggagaaga
     1861 aagatctcca agaagtgctt caatcagact ctaatcatgt tccttcggta ttgacagaac
     1921 ttaggagtgg attagatcaa acaattcatt atttcaacgc agttcaaagt tataagtcaa
     1981 aggtagattc ggcaacatcg attttggaag ccttggctga ggagttaaac aaaaaccagt
     2041 tgactaatca aaggcatgaa gccgcgcatg agttaatgag ggcgcaaacc ggcctggaga
     2101 ttccgcaatt agctcaagaa ctcgtgcagc ttgaaagaca ttacacacat tttgctaaag
     2161 cgtataccgc tctccttcag gaaattcaca gacgacaaac ctatgaaaat tgtgtaagat
     2221 ccattgtgga cgaatttgtg ggacgcctgg agaaggaaca acaggcggag gcgaaatgtc
     2281 gtatcgattt tttcaatcaa tatggtgatt atttacccca gactctatgg ggagctgtaa
     2341 cagatcctcc tttacatttt gaaattatag agcaccaata tacagaattg ccaaacgtaa
     2401 aggtaattcc tgataaaaac gataagaaat cgaagcaacg tgaaaaatct tctactacag
     2461 cctctaagcg atgactcgag tcatgtaatt agttatgtca cgcttacatt cacgccctcc
     2521 ccccacatcc gctctaaccg aaaaggaagg agttagacaa cctgaagtct aggtccctat
     2581 ttattttttt atagttatgt tagtattaag aacgttattt atatttcaaa tttttctttt
     2641 ttttctgtac agacgcgtgt acgcatgtaa cattatactg aaaaccttgc ttgagaaggt
     2701 tttgggacgc tcgaaggctt taatttgcgg ccggtaccca attcgcccta tagtgagtcg
     2761 tattacgcgc gctcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt
     2821 acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag
     2881 gcccgcaccg atcgcccttc ccaacagttg cgcagcctga atggcgaatg gacgcgccct
     2941 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
     3001 ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
     3061 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac
     3121 ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct
     3181 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt
     3241 tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta taagggattt
     3301 tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
     3361 ttaacaaaat attaacgctt acaatttcct gatgcggtat tttctcctta cgcatctgtg
     3421 cggtatttca caccgcatag ggtaataact gatataatta aattgaagct ctaatttgtg
     3481 agtttagtat acatgcattt acttataata cagtttttta gttttgctgg ccgcatcttc
     3541 tcaaatatgc ttcccagcct gcttttctgt aacgttcacc ctctacctta gcatcccttc
     3601 cctttgcaaa tagtcctctt ccaacaataa taatgtcaga tcctgtagag accacatcat
     3661 ccacggttct atactgttga cccaatgcgt ctcccttgtc atctaaaccc acaccgggtg
     3721 tcataatcaa ccaatcgtaa ccttcatctc ttccacccat gtctctttga gcaataaagc
     3781 cgataacaaa atctttgtcg ctcttcgcaa tgtcaacagt acccttagta tattctccag
     3841 tagataggga gcccttgcat gacaattctg ctaacatcaa aaggcctcta ggttcctttg
     3901 ttacttcttc tgccgcctgc ttcaaaccgc taacaatacc tgggcccacc acaccgtgtg
     3961 cattcgtaat gtctgcccat tctgctattc tgtatacacc cgcagagtac tgcaatttga
     4021 ctgtattacc aatgtcagca aattttctgt cttcgaagag taaaaaattg tacttggcgg
     4081 ataatgcctt tagcggctta actgtgccct ccatggaaaa atcagtcaag atatccacat
     4141 gtgtttttag taaacaaatt ttgggaccta atgcttcaac taactccagt aattccttgg
     4201 tggtacgaac atccaatgaa gcacacaagt ttgtttgctt ttcgtgcatg atattaaata
     4261 gcttggcagc aacaggacta ggatgagtag cagcacgttc cttatatgta gctttcgaca
     4321 tgatttatct tcgtttcctg caggtttttg ttctgtgcag ttgggttaag aatactgggc
     4381 aatttcatgt ttcttcaaca ctacatatgc gtatatatac caatctaagt ctgtgctcct
     4441 tccttcgttc ttccttctgt tcggagatta ccgaatcaaa aaaatttcaa ggaaaccgaa
     4501 atcaaaaaaa agaataaaaa aaaaatgatg aattgaaaag gtggtatggt gcactctcag
     4561 tacaatctgc tctgatgccg catagttaag ccagccccga cacccgccaa cacccgctga
     4621 cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc
     4681 cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga gacgaaaggg
     4741 cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt cttagacgga
     4801 tcgcttgcct gtaacttaca cgcgcctcgt atcttttaat gatggaataa tttgggaatt
     4861 tactctgtgt ttatttattt ttatgttttg tatttggatt ttagaaagta aataaagaag
     4921 gtagaagagt tacggaatga agaaaaaaaa ataaacaaag gtttaaaaaa tttcaacaaa
     4981 aagcgtactt tacatatata tttattagac aagaaaagca gattaaatag atatacattc
     5041 gattaacgat aagtaaaatg taaaatcaca ggattttcgt gtgtggtctt ctacacagac
     5101 aagatgaaac aattcggcat taatacctga gagcaggaag agcaagataa aaggtagtat
     5161 ttgttggcga tccccctaga gtcttttaca tcttcggaaa acaaaaacta ttttttcttt
     5221 aatttctttt tttactttct atttttaatt tatatattta tattaaaaaa tttaaattat
     5281 aattattttt atagcacgtg atgaaaagga cccaggtggc acttttcggg gaaatgtgcg
     5341 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     5401 ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt
     5461 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     5521 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga
     5581 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat
     5641 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca
     5701 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt
     5761 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     5821 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct
     5881 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     5941 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac
     6001 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat
     6061 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     6121 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     6181 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     6241 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     6301 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
     6361 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     6421 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     6481 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     6541 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     6601 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa
     6661 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     6721 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     6781 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     6841 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     6901 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     6961 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     7021 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     7081 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     7141 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     7201 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagag
//
