LOCUS       sequence-417181-        5643 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Mammalian expression of ATG101.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5643)
  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 5643)
  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-ATG101
FEATURES             Location/Qualifiers
     source          1..5643
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        4..383
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        385..661
                     /label=chicken beta-actin promoter
     intron          662..1678
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1602..1624
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1686..1705
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1752..1761
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1767..1790
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1827..1850
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1857..1880
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(2664..2683)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2729..2784
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2783..2802)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3145..3161)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3145..3161)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3158..3180)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3169..3185
                     /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(3193..3199,3200..3217,3218..3223))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3238..3259
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3318..3513
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3364..3499
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3519..3653
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3556..3575)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3610..3629
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3721..3738)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3892..4480)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3972..3991)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4651..5442,5443..5511))
                     /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     5274..5293
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5512..5616)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
       61 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      121 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      181 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac
      241 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
      301 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
      361 tattagtcat cgctattacc atggtcgagg tgagccccac gttctgcttc actctcccca
      421 tctccccccc ctccccaccc ccaattttgt atttatttat tttttaatta ttttgtgcag
      481 cgatgggggc gggggggggg gggggsgcgc gccaggsggg gggggggggg gggaggggsg
      541 gggsggggsg aggsggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgcgctgcc ttcgccccgt gccccgctcc gccgccgcct cgcgccgccc
      721 gccccggctc tgactgaccg cgttactccc acaggtgagc gggcgggacg gcccttctcc
      781 tccgggctgt aattagcgct tggtttaatg acggcttgtt tcttttctgt ggctgcgtga
      841 aagccttgag gggctccggg agggcccttt gtgcgggggg agcggctcgg ggggtgcgtg
      901 cgtgtgtgtg tgcgtgggga gcgccgcgtg cggctccgcg ctgcccggcg gctgtgagcg
      961 ctgcgggcgc ggcgcggggc tttgtgcgct ccgcagtgtg cgcgagggga gcgcggccgg
     1021 gggcggtgcc ccgcggtgcg gggggggctg cgaggggaac aaaggctgcg tgcggggtgt
     1081 gtgcgtgggg gggtgagcag ggggtgtggg cgcgtcggtc gggctgcaac cccccctgca
     1141 cccccctccc cgagttgctg agcacggccc ggcttcgggt gcggggctcc gtacggggcg
     1201 tggcgcgggg ctcgccgtgc cgggcggggg gtggcggcag gtgggggtgc cgggcggggc
     1261 ggggccgcct cgggccgggg rgggctcggg ggaggggcgc ggcggccccc ggagcgccgg
     1321 cggctgtcga ggcgcggcga gccgcagcca ttgcctttta tggtaatcgt gcgagagggc
     1381 gcagggactt cctttgtccc aaatctgtgc ggagccgaaa tctgggaggc gccgccgcac
     1441 cccctctagc gggcgcgggg cgaagcggtg cggcgccggc aggaaggaaa tgggcgggga
     1501 gggccttcgt gcgtcgccgc gccgccgtcc ccttctccct ctccagcctc ggggctgtcc
     1561 gcggggggac ggctgccttc gggggggacg gggcagggcg gggttcggct tctggcgtgt
     1621 gaccggcggc tctagagcct ctgctaacca tgttcatgcc ttcttctttt tcctacagct
     1681 cctgggcaac gtgctggtta ttgtgctgtc tcatcatttt ggcaaagaat tcctcgatcg
     1741 acggtatcga tgccaccatg gctagctgga gccacccgca gttcgagaaa ggtggaggtg
     1801 cccgaggtgg atcgggaggt ggatcgtgga gccacccgca gttcgaaaaa ggattcgatt
     1861 acaaggatga cgatgacaag ggtaccatga attgccggtc agaggtcctg gaagtgagcg
     1921 tggaaggtag acaggtggaa gaggctatgc tggccgtgct gcatacagtg ctgctgcaca
     1981 gatcaactgg caagttccat tacaagaaag aaggaacata ttccatcgga accgtgggta
     2041 ctcaggacgt cgattgcgac ttcattgatt ttacttacgt gcgcgtcagc tccgaggaac
     2101 tggacagagc actccgtaag gtggtcggag aattcaaaga tgccctgagg aacagtggcg
     2161 gagacggtct gggccagatg agcctcgagt tctatcagaa gaaaaagtct agatggccat
     2221 tttcagatga atgtatcccc tgggaagtgt ggaccgtgaa ggtccacgtg gtcgccctgg
     2281 ctactgagca ggaacggcag atctgccgcg agaaagtggg cgaaaagctc tgtgagaaga
     2341 tcatcaacat cgtggaagtc atgaaccgtc atgagtacct gcccaagatg cctacacaga
     2401 gcgaggtgga taatgtcttc gacaccggtc tgcgggatgt gcagccctac ctctataaga
     2461 tctcctttca gattacagac gctctgggca cctctgtcac cactacaatg cgcaggctca
     2521 tcaaagacac cctggcactc taaggtaccc ccgggtgata tcttaattaa ttaactcgag
     2581 gaattcactc ctcaggtgca ggctgcctat cagaaggtgg tggctggtgt ggccaatgcc
     2641 ctggctcaca aataccactg agatcttttt ccctctgcca aaaattatgg ggacatcatg
     2701 aagccccttg agcatctgac ttctggctaa taaaggaaat ttattttcat tgcaatagtg
     2761 tgttggaatt ttttgtgtct ctcactcgga aggacatatg ggagggcaaa tcatttaaaa
     2821 catcagaatg agtatttggt ttagagtttg gcaacatatg cccatatgct ggctgccatg
     2881 aacaaaggtt ggctataaag aggtcatcag tatatgaaac agccccctgc tgtccattcc
     2941 ttattccata gaaaagcctt gacttgaggt tagatttttt ttatattttg ttttgtgtta
     3001 tttttttctt taacatccct aaaattttcc ttacatgttt tactagccag atttttcctc
     3061 ctctcctgac tactcccagt catagctgtc cctcttctct tatggagatc cctcgacctg
     3121 cagcccaagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct
     3181 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg
     3241 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     3301 gtcgtgccag cggatccgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     3361 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     3421 ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga
     3481 ggaggctttt ttggaggcct aggcttttgc aaaaagctaa cttgtttatt gcagcttata
     3541 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     3601 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgg atccgctgca
     3661 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     3721 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     3781 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     3841 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     3901 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     3961 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     4021 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     4081 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     4141 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     4201 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     4261 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     4321 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     4381 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     4441 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     4501 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     4561 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     4621 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     4681 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     4741 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
     4801 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     4861 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     4921 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     4981 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     5041 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     5101 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     5161 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     5221 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     5281 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     5341 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     5401 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     5461 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     5521 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     5581 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     5641 tgg
//
