LOCUS       sequence-400872-        5846 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Mammalian expression of ATG13 C-terminal and ULK1 MIT domain fusion 
            complex.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5846)
  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 5846)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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-TSF-ATG13 (450-517)-ULK1 (836-1050)
FEATURES             Location/Qualifiers
     source          1..5846
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(4..108)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        139..518
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        520..795
                     /label=chicken beta-actin promoter
     intron          796..1812
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1736..1758
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1820..1839
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1887..1896
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1902..1925
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1962..1985
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1992..2015
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2019..2039
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(3002..3021)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3067..3122
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3121..3140)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3483..3499)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3483..3499)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3496..3518)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3507..3523
                     /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(3531..3537,3538..3555,3556..3561))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3576..3597
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3656..3851
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3702..3837
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3857..3991
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3894..3913)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3948..3967
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4059..4076)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4230..4818)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4310..4329)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4989..5780,5781..5846,1..3))
                     /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     5612..5631
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
       61 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
      121 aaaagtgcca cctgggtcga cattgattat tgactagtta ttaatagtaa tcaattacgg
      181 ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc
      241 cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca
      301 tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg
      361 cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg
      421 acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt
      481 ggcagtacat ctacgtatta gtcatcgcta ttaccatggt cgaggtgagc cccacgttct
      541 gcttcactct ccccatctcc cccccctccc cacccccaat tttgtattta tttatttttt
      601 aattattttg tgcagcgatg ggggcggggg gggggggggg gcgcgccagg cggggcgggg
      661 cggggcgagg ggcggggcgg ggcgaggcgg agaggtgcgg cggcagccaa tcagagcggc
      721 gcgctccgaa agtttccttt tatggcgagg cggcggcggc ggcggcccta taaaaagcga
      781 agcgcgcggc gggcgggagt cgctgcgcgc tgccttcgcc ccgtgccccg ctccgccgcc
      841 gcctcgcgcc gcccgccccg gctctgactg accgcgttac tcccacaggt gagcgggcgg
      901 gacggccctt ctcctccggg ctgtaattag cgcttggttt aatgacggct tgtttctttt
      961 ctgtggctgc gtgaaagcct tgaggggctc cgggagggcc ctttgtgcgg ggggagcggc
     1021 tcggggggtg cgtgcgtgtg tgtgtgcgtg gggagcgccg cgtgcggctc cgcgctgccc
     1081 ggcggctgtg agcgctgcgg gcgcggcgcg gggctttgtg cgctccgcag tgtgcgcgag
     1141 gggagcgcgg ccgggggcgg tgccccgcgg tgcggggggg gctgcgaggg gaacaaaggc
     1201 tgcgtgcggg gtgtgtgcgt gggggggtga gcagggggtg tgggcgcgtc ggtcgggctg
     1261 caaccccccc tgcacccccc tccccgagtt gctgagcacg gcccggcttc gggtgcgggg
     1321 ctccgtacgg ggcgtggcgc ggggctcgcc gtgccgggcg gggggtggcg gcaggtgggg
     1381 gtgccgggcg gggcggggcc gcctcgggcc ggggagggct cgggggaggg gcgcggcggc
     1441 ccccggagcg ccggcggctg tcgaggcgcg gcgagccgca gccattgcct tttatggtaa
     1501 tcgtgcgaga gggcgcaggg acttcctttg tcccaaatct gtgcggagcc gaaatctggg
     1561 aggcgccgcc gcaccccctc tagcgggcgc ggggcgaagc ggtgcggcgc cggcaggaag
     1621 gaaatgggcg gggagggcct tcgtgcgtcg ccgcgccgcc gtccccttct ccctctccag
     1681 cctcggggct gtccgcgggg ggacggctgc cttcgggggg gacggggcag ggcggggttc
     1741 ggcttctggc gtgtgaccgg cggctctaga gcctctgcta accatgttca tgccttcttc
     1801 tttttcctac agctcctggg caacgtgctg gttattgtgc tgtctcatca ttttggcaaa
     1861 gaattcctcg atcgacggta tcgatagcca ccatggctag ctggagccac ccgcagttcg
     1921 agaaaggtgg aggtgcccga ggtggatcgg gaggtggatc gtggagccac ccgcagttcg
     1981 aaaaaggatt cgattacaag gatgacgatg acaagaccga aaacctgtat tttcagggtg
     2041 gtaccaaacc cgctttttcc aaggacgata tcctccctat ggatctggga accttctaca
     2101 gggagtttca gaatccccct cagctcagta gcctgagcat tgacatcggt gcccagtcca
     2161 tggctgagga cctcgattct ctgcctgaaa aactggctgt gcacgagaag aacgtcagag
     2221 aattcgacgc atttgtggag accctgcagg gtagcgacga agcgatggag caggaacaca
     2281 ctgagatcct gaggggtctc agattcaccc tgctctttgt gcagcatgtc ctggaaattg
     2341 ctgcactcaa gggtagtgct agcgaggcag ctggtggacc agaataccag ctgcaggaga
     2401 gcgtggtcgc cgaccagatc tctctgctct caagggaatg gggcttcgct gagcagctgg
     2461 tcctctatct gaaggtggcc gaactgctca gtagcggcct gcagtccgcc atcgatcaga
     2521 ttagagctgg aaagctctgc ctgtcctcta ctgtgaaaca ggtggtccgg cgcctgaacg
     2581 agctctacaa ggccagtgtg gtcagctgtc agggactctc tctgcgtctc cagcggttct
     2641 ttctggacaa acagcgcctg ctcgatagga tccactcaat tacagctgaa cgcctgatct
     2701 tcagccatgc agtgcagatg gtccagtccg cagccctgga cgaaatgttt cagcacaggg
     2761 agggatgcgt ccccagatac cataaggctc tgctcctgct cgagggtctg cagcacatgc
     2821 tcagcgacca ggccgatatc gaaaatgtga ctaagtgcaa actgtgtatt gagaggagac
     2881 tctccgctct gctcacaggc atctgtgcat gactcgagga attcactcct caggtgcagg
     2941 ctgcctatca gaaggtggtg gctggtgtgg ccaatgccct ggctcacaaa taccactgag
     3001 atctttttcc ctctgccaaa aattatgggg acatcatgaa gccccttgag catctgactt
     3061 ctggctaata aaggaaattt attttcattg caatagtgtg ttggaatttt ttgtgtctct
     3121 cactcggaag gacatatggg agggcaaatc atttaaaaca tcagaatgag tatttggttt
     3181 agagtttggc aacatatgcc catatgctgg ctgccatgaa caaaggttgg ctataaagag
     3241 gtcatcagta tatgaaacag ccccctgctg tccattcctt attccataga aaagccttga
     3301 cttgaggtta gatttttttt atattttgtt ttgtgttatt tttttcttta acatccctaa
     3361 aattttcctt acatgtttta ctagccagat ttttcctcct ctcctgacta ctcccagtca
     3421 tagctgtccc tcttctctta tggagatccc tcgacctgca gcccaagctt ggcgtaatca
     3481 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     3541 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     3601 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagcg gatccgcatc
     3661 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     3721 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     3781 aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     3841 gcttttgcaa aaagctaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     3901 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     3961 actcatcaat gtatcttatc atgtctggat ccgctgcatt aatgaatcgg ccaacgcgcg
     4021 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     4081 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     4141 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     4201 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     4261 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     4321 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     4381 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     4441 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     4501 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     4561 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     4621 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     4681 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     4741 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     4801 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     4861 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     4921 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     4981 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     5041 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     5101 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     5161 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     5221 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     5281 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     5341 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     5401 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     5461 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     5521 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     5581 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     5641 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     5701 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     5761 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     5821 agggcgacac ggaaatgttg aatact
//
