LOCUS       sequence-400962-        7016 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of GST tagged ATG14.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7016)
  AUTHORS   Chen M, Ren X, Nguyen TN, Khuu G, Cook ASI, Lazarou M, Hurley JH
  TITLE     Structure and activation of the human autophagy-initiating 
            ULK1C:PI3KC3-C1 supercomplex
  JOURNAL   bioRxiv 2023.06.01.543278
REFERENCE   2  (bases 1 to 7016)
  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: "bioRxiv 
            2023.06.01.543278"
COMMENT     Sequence Label: pCAG-GST-ATG14
FEATURES             Location/Qualifiers
     source          1..7016
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        382..657
                     /label=chicken beta-actin promoter
     intron          658..1674
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1598..1620
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1682..1701
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1749..1758
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1755..2411
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MASPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNK
                     KFELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIR
                     YGVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVV
                     LYMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2369..2391
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2439..2459
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(4034..4053)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4099..4154
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4153..4172)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(4515..4531)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4515..4531)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4528..4550)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4539..4555
                     /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(4563..4569,4570..4587,4588..4593))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4608..4629
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4688..4883
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4734..4869
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4889..5023
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4926..4945)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4980..4999
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5091..5108)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5262..5850)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5342..5361)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6021..6812,6813..6881))
                     /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     6644..6663
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6882..6986)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtcgaggtga gccccacgtt ctgcttcact ctccccatct
      421 cccccccctc cccaccccca attttgtatt tatttatttt ttaattattt tgtgcagcga
      481 tgggggcggg gggggggggg gsgcgcgcca ggcggggcgg ggcggggcga ggggcggggc
      541 ggggcgaggc ggagaggtgc ggcggcagcc aatcagagcg gcgcgctccg aaagtttcct
      601 tttatggcga ggcggcggcg gcggcggccc tataaaaagc gaagcgcgcg gcgggcggga
      661 gtcgctgcgc gctgccttcg ccccgtgccc cgctccgccg ccgcctcgcg ccgcccgccc
      721 cggctctgac tgaccgcgtt actcccacag gtgagcgggc gggacggccc ttctcctccg
      781 ggctgtaatt agcgcttggt ttaatgacgg cttgtttctt ttctgtggct gcgtgaaagc
      841 cttgaggggc tccgggaggg ccctttgtgc ggggggagcg gctcgggggg tgcgtgcgtg
      901 tgtgtgtgcg tggggagcgc cgcgtgcggc tccgcgctgc ccggcggctg tgagcgctgc
      961 gggcgcggcg cggggctttg tgcgctccgc agtgtgcgcg aggggagcgc ggccgggggc
     1021 ggtgccccgc ggtgcggggg gggctgcgag gggaacaaag gctgcgtgcg gggtgtgtgc
     1081 gtgggggggt gagcaggggg tgtgggcgcg tcggtcgggc tgcaaccccc cctgcacccc
     1141 cctccccgag ttgctgagca cggcccggct tcgggtgcgg ggctccgtac ggggcgtggc
     1201 gcggggctcg ccgtgccggg cggggggtgg cggcaggtgg gggtgccggg cggggcgggg
     1261 ccgcctcggg ccggggaggg ctcgggggag gggcgcggcg gcccccggag cgccggcggc
     1321 tgtcgaggcg cggcgagccg cagccattgc cttttatggt aatcgtgcga gagggcgcag
     1381 ggacttcctt tgtcccaaat ctgtgcggag ccgaaatctg ggaggcgccg ccgcaccccc
     1441 tctagcgggc gcggggcgaa gcggtgcggc gccggcagga aggaaatggg cggggagggc
     1501 cttcgtgcgt cgccgcgccg ccgtcccctt ctccctctcc agcctcgggg ctgtccgcgg
     1561 ggggacggct gccttcgggg gggacggggc agggcggggt tcggcttctg gcgtgtgacc
     1621 ggcggctcta gagcctctgc taaccatgtt catgccttct tctttttcct acagctcctg
     1681 ggcaacgtgc tggttattgt gctgtctcat cattttggca aagaattcct cgatcgacgg
     1741 tatcgatagc caccatggca tcccctatac taggttattg gaaaattaag ggccttgtgc
     1801 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc
     1861 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc
     1921 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata
     1981 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc
     2041 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact
     2101 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag
     2161 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt
     2221 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa
     2281 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat
     2341 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc
     2401 atcctccaaa atcggatgat tacgatatcc caacgaccga aaacctgtat tttcagggtg
     2461 gtaccatggc ttccccatct ggaaagggag ctagggctct ggaggctcct ggatgcggac
     2521 cacggcccct ggccagagac ctggtggata gcgtggacga tgctgagggg ctgtacgtgg
     2581 ccgtggaaag atgccccctg tgtaatacca cacggagaag gctgacatgc gccaagtgtg
     2641 tgcagtccgg ggatttcgtg tattttgacg gacgcgatcg ggagagattc attgacaaga
     2701 aagaaaggct gtcccgcctg aagtctaaac aggaggaatt tcagaaagag gtgctgaagg
     2761 ccatggaggg caagtggatc actgatcagc tgaggtggaa aatcatgtcc tgcaagatgc
     2821 gcattgagca gctgaaacag accatctgta agggaaacga ggaaatggag aaaaatagcg
     2881 aaggcctgct gaagacaaaa gaaaagaatc agaagctgta ctcccgggct cagagacacc
     2941 aggagaagaa agaaaaaatt cagaggcata accgcaagct gggcgatctg gtggagaaga
     3001 aaaccatcga cctgaggtct cactatgaac gcctggccaa tctgcgccgg agccatatcc
     3061 tggagctgac atccgtgatc ttcccaattg aggaagtgaa aactggagtg agggaccccg
     3121 ctgatgtgag ctccgagagt gactcagcca tgacctctag tacagtgagt aagctggctg
     3181 aagccagaag gactacctac ctgtcaggcc gctgggtgtg cgacgatcac aacggcgata
     3241 caagcatctc cattactggg ccctggatta gcctgcctaa caatggcgac tatagtgcct
     3301 actattcatg ggtggaggaa aagaaaacaa ctcaggggcc agatatggag cagtctaatc
     3361 ccgcttacac tatcagtgcc gctctgtgct atgccaccca gctggtgaac atcctgtctc
     3421 atattctgga cgtgaacctg cccaagaaac tgtgcaattc agagttctgt ggcgaaaacc
     3481 tgagcaaaca gaagtttact agggctgtga agaaactgaa cgccaatatt ctgtacctgt
     3541 gttttagcca gcacgtgaat ctggatcagc tgcagcctct gcataccctg cgcaacctga
     3601 tgtatctggt gtccccttca agcgagcacc tgggaaggtc tggaccattc gaagtgagag
     3661 ctgatctgga ggaaagcatg gagtttgtgg accctggagt ggccggcgag tctgacgaaa
     3721 gtggggatga gcgggtgtct gacgaggaaa ctgatctggg aaccgactgg gaaaacctgc
     3781 ctagtccaag attctgtgac atcccatctc agagtgtgga ggtgtcacag agccagtcca
     3841 cacaggcttc cccccctatt gcttcctcta gtgctggagg aatgatctca agcgccgctg
     3901 cctcagtgac cagctggttt aaggcctaca caggccatag gtgactcgag gaattcactc
     3961 ctcaggtgca ggctgcctat cagaaggtgg tggctggtgt ggccaatgcc ctggctcaca
     4021 aataccactg agatcttttt ccctctgcca aaaattatgg ggacatcatg aagccccttg
     4081 agcatctgac ttctggctaa taaaggaaat ttattttcat tgcaatagtg tgttggaatt
     4141 ttttgtgtct ctcactcgga aggacatatg ggagggcaaa tcatttaaaa catcagaatg
     4201 agtatttggt ttagagtttg gcaacatatg cccatatgct ggctgccatg aacaaaggtt
     4261 ggctataaag aggtcatcag tatatgaaac agccccctgc tgtccattcc ttattccata
     4321 gaaaagcctt gacttgaggt tagatttttt ttatattttg ttttgtgtta tttttttctt
     4381 taacatccct aaaattttcc ttacatgttt tactagccag atttttcctc ctctcctgac
     4441 tactcccagt catagctgtc cctcttctct tatggagatc cctcgacctg cagcccaagc
     4501 ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
     4561 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
     4621 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     4681 cggatccgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc
     4741 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt
     4801 atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt
     4861 ttggaggcct aggcttttgc aaaaagctaa cttgtttatt gcagcttata atggttacaa
     4921 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     4981 tggtttgtcc aaactcatca atgtatctta tcatgtctgg atccgctgca ttaatgaatc
     5041 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     5101 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     5161 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     5221 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     5281 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     5341 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     5401 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     5461 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     5521 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     5581 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     5641 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     5701 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     5761 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     5821 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     5881 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     5941 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     6001 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     6061 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     6121 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     6181 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     6241 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     6301 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     6361 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     6421 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     6481 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     6541 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     6601 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     6661 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     6721 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     6781 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     6841 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     6901 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     6961 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgggtc
//
