LOCUS       sequence-236911-        7300 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expressing GFP-Atg17.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7300)
  AUTHORS   Liu X, Mao K, Yu AYH, Omairi-Nasser A, Austin J 2nd, Glick BS, Yip 
            CK, Klionsky DJ
  TITLE     The Atg17-Atg31-Atg29 Complex Coordinates with Atg11 to Recruit the 
            Vam7 SNARE and Mediate Autophagosome-Vacuole Fusion.
  JOURNAL   Curr Biol. 2016 Jan 25;26(2):150-160. doi: 
            10.1016/j.cub.2015.11.054. Epub 2016 Jan 7.
  PUBMED    26774783
REFERENCE   2  (bases 1 to 7300)
  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"; doi: 
            "10.1016/j.cub.2015.11.054"; journalName: "Curr Biol"; date: 
            "2016-01-25- 25"; volume: "26"; issue: "2"; pages: "150-160"
COMMENT     Sequence Label: pCu-GFP-Atg17(416)
FEATURES             Location/Qualifiers
     source          1..7300
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    36..57
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(72..77,78..95,96..102)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    110..126
                     /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     115..137
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     134..150
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     134..150
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     169..189
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        171..189
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        208..442
                     /gene="S. cerevisiae CUP1"
                     /label=CUP1 promoter
                     /note="inducible metallothionein promoter, regulated by 
                     Cu2+"
     CDS             450..1163
                     /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(480..508)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     1103..1123
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     RBS             2246..2257
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     terminator      2466..2713
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(2482..2500)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(2731..2750)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2732..2750)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2760..2777)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2760..2776)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2769..2791)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2917..3372
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3004..3023)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3214..3235
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3503..4306)
                     /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(4307..4522)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(4546..4565)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4665..4687
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4725..4743)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    4784..5287
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        5324..5428
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5429..5497,5498..6289)
                     /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(5647..5666)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6460..7048
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6949..6968
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7202..7219
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct cactcattag
       61 gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga
      121 taacaatttc acacaggaaa cagctatgac catgattacg ccaagcgcgc aattaaccct
      181 cactaaaggg aacaaaagct ggagctccta gttagaaaaa gacatttttg ctgtcagtca
      241 ctgtcaagag attcttttgc tggcatttct tctagaagca aaaagagcga tgcgtctttt
      301 ccgctgaacc gttccagcaa aaaagactac caacgcaata tggattgtca gaatcatata
      361 aaagagaagc aaataactcc ttgtcttgta tcaattgcat tataatatct tcttgttagt
      421 gcaatatcat atagaagtca tcgactagta tgagtaaagg agaagaactt ttcactggag
      481 ttgtcccaat tcttgttgaa ttagatggtg atgttaatgg gcacaaattt tctgtcagtg
      541 gagagggtga aggtgatgca acatacggaa aacttaccct taaatttatt tgcactactg
      601 gaaaactacc tgttccatgg ccaacacttg tcactacttt cacttatggt gttcaatgct
      661 tttcaagata cccagatcat atgaaacggc atgacttttt caagagtgcc atgcccgaag
      721 gttatgtaca ggaaagaact atatttttca aagatgacgg gaactacaag acacgtgctg
      781 aagtcaagtt tgaaggtgat acccttgtta atagaatcga gttaaaaggt attgatttta
      841 aagaagatgg aaacattctt ggacacaaat tggaatacaa ctataactca cacaatgtat
      901 acatcatggc agacaaacaa aagaatggaa tcaaagttaa cttcaaaatt agacacaaca
      961 ttgaagatgg aagcgttcaa ctagcagacc attatcaaca aaatactcca attggcgatg
     1021 gccctgtcct tttaccagac aaccattacc tgtccacaca atctgccctt tcgaaagatc
     1081 ccaacgaaaa gagagaccac atggtccttc ttgagtttgt aacagctgct gggattacac
     1141 atggcatgga tgaactatac aaacccgggc tgcaggaatt cgatatcaag cttatcgata
     1201 tgaacgaagc agatgttaca aaatttgtta ataatgccag gaaaacgctg accgatgctc
     1261 aacttttatg ttcaagtgct aatttaagga ttgtagatat taagaaaaaa ttgtcatctt
     1321 ggcagttgag tatttcaaaa ctcaattttc taatagttgg cttgagacag caaggaaagt
     1381 ttctttacac tattttaaag gaaggcattg ggacaaagct aatccagaaa caatggaatc
     1441 aagctgtctt agtggtctta gttgatgaga tgaagtactg gcaatatgaa atcacttcta
     1501 aagtacaaag gttggatggt atagtaaatg aattgagcat atcagaaaaa gatgataccg
     1561 atccctctaa actaggagat tacatctcga gagacaacgt taacttattg aatgacaaac
     1621 tgaaagaagt gccagtaatt gagcgtcaaa ttgaaaacat taagcttcaa tacgaaaata
     1681 tggtcagaaa agttaacaaa gaactgattg ataccaagtt gacggacgta actcaaaaat
     1741 tccaaagtaa atttggtata gataacctga tggaaacaaa tgttgcagag cagtttagca
     1801 gggaactaac ggaccttgaa aaagatttag cagagataat gaattcattg acacagcact
     1861 ttgataagac attgctgtta caagataaaa aaattgataa cgatgaacgt gaggagctgt
     1921 ttaaggtggt acaaggcgac gacaaagaac tatacaacat tttcaaaact ctgcatgggg
     1981 taattgatga cgtggacaaa acaattctta acttgggtca atttttgcag gcaaaaataa
     2041 aggaaaagac agaactacac agcgaagttt ctgaaataat aaacgatttc aatagaaatt
     2101 tggaatatct attaatcttt aaagatattt ccaatctgat tgatagcttc aaaaattcct
     2161 gtacacaaga tattcaaaca actaaggaac tttgtgaatt ttatgataat tttgaagaaa
     2221 gctacggtaa cttagttcta gaagcaaaga ggagaaagga tgtggcaaac agaatgaaaa
     2281 ctatattgaa agattgtgaa aagcagttgc agaatttaga tgctcaggac caggaagaac
     2341 gtcagaattt tatagcggaa aatggaactt atcttcctga gacaatctgg cccggtaaaa
     2401 ttgacgattt ttcttccctg tacactttaa attacaacgt gaagaatcct taggtcgacc
     2461 tcgagtcatg taattagtta tgtcacgctt acattcacgc cctcccccca catccgctct
     2521 aaccgaaaag gaaggagtta gacaacctga agtctaggtc cctatttatt tttttatagt
     2581 tatgttagta ttaagaacgt tatttatatt tcaaattttt cttttttttc tgtacagacg
     2641 cgtgtacgca tgtaacatta tactgaaaac cttgcttgag aaggttttgg gacgctcgaa
     2701 ggctttaatt tgcggccggt acccaattcg ccctatagtg agtcgtatta cgcgcgctca
     2761 ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc
     2821 cttgcagcac atcccccttt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc
     2881 ccttcccaac agttgcgcag cctgaatggc gaatggacgc gccctgtagc ggcgcattaa
     2941 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc
     3001 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     3061 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     3121 aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     3181 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     3241 cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct
     3301 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa
     3361 cgcttacaat ttcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     3421 catagggtaa taactgatat aattaaattg aagctctaat ttgtgagttt agtatacatg
     3481 catttactta taatacagtt ttttagtttt gctggccgca tcttctcaaa tatgcttccc
     3541 agcctgcttt tctgtaacgt tcaccctcta ccttagcatc ccttcccttt gcaaatagtc
     3601 ctcttccaac aataataatg tcagatcctg tagagaccac atcatccacg gttctatact
     3661 gttgacccaa tgcgtctccc ttgtcatcta aacccacacc gggtgtcata atcaaccaat
     3721 cgtaaccttc atctcttcca cccatgtctc tttgagcaat aaagccgata acaaaatctt
     3781 tgtcgctctt cgcaatgtca acagtaccct tagtatattc tccagtagat agggagccct
     3841 tgcatgacaa ttctgctaac atcaaaaggc ctctaggttc ctttgttact tcttctgccg
     3901 cctgcttcaa accgctaaca atacctgggc ccaccacacc gtgtgcattc gtaatgtctg
     3961 cccattctgc tattctgtat acacccgcag agtactgcaa tttgactgta ttaccaatgt
     4021 cagcaaattt tctgtcttcg aagagtaaaa aattgtactt ggcggataat gcctttagcg
     4081 gcttaactgt gccctccatg gaaaaatcag tcaagatatc cacatgtgtt tttagtaaac
     4141 aaattttggg acctaatgct tcaactaact ccagtaattc cttggtggta cgaacatcca
     4201 atgaagcaca caagtttgtt tgcttttcgt gcatgatatt aaatagcttg gcagcaacag
     4261 gactaggatg agtagcagca cgttccttat atgtagcttt cgacatgatt tatcttcgtt
     4321 tcctgcaggt ttttgttctg tgcagttggg ttaagaatac tgggcaattt catgtttctt
     4381 caacactaca tatgcgtata tataccaatc taagtctgtg ctccttcctt cgttcttcct
     4441 tctgttcgga gattaccgaa tcaaaaaaat ttcaaggaaa ccgaaatcaa aaaaaagaat
     4501 aaaaaaaaaa tgatgaattg aaaaggtggt atggtgcact ctcagtacaa tctgctctga
     4561 tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc
     4621 ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg
     4681 tcagaggttt tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct
     4741 atttttatag gttaatgtca tgataataat ggtttcttag acggatcgct tgcctgtaac
     4801 ttacacgcgc ctcgtatctt ttaatgatgg aataatttgg gaatttactc tgtgtttatt
     4861 tatttttatg ttttgtattt ggattttaga aagtaaataa agaaggtaga agagttacgg
     4921 aatgaagaaa aaaaaataaa caaaggttta aaaaatttca acaaaaagcg tactttacat
     4981 atatatttat tagacaagaa aagcagatta aatagatata cattcgatta acgataagta
     5041 aaatgtaaaa tcacaggatt ttcgtgtgtg gtcttctaca cagacaagat gaaacaattc
     5101 ggcattaata cctgagagca ggaagagcaa gataaaaggt agtatttgtt ggcgatcccc
     5161 ctagagtctt ttacatcttc ggaaaacaaa aactattttt tctttaattt ctttttttac
     5221 tttctatttt taatttatat atttatatta aaaaatttaa attataatta tttttatagc
     5281 acgtgatgaa aaggacccag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     5341 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     5401 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     5461 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     5521 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     5581 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     5641 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
     5701 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     5761 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     5821 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     5881 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     5941 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact
     6001 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     6061 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     6121 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     6181 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     6241 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     6301 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     6361 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     6421 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     6481 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     6541 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     6601 tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     6661 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     6721 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     6781 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     6841 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     6901 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     6961 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     7021 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     7081 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga
     7141 taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg
     7201 cagcgagtca gtgagcgagg aagcggaaga gcgcccaata cgcaaaccgc ctctccccgc
     7261 gcgttggccg attcattaat gcagctggca cgacaggttt
//
