LOCUS       sequence-212738-        7352 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of hATG5.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7352)
  AUTHORS   Kim M, Sandford E, Gatica D, Qiu Y, Liu X, Zheng Y, Schulman BA, Xu 
            J, Semple I, Ro SH, Kim B, Mavioglu RN, Tolun A, Jipa A, Takats S, 
            Karpati M, Li JZ, Yapici Z, Juhasz G, Lee JH, Klionsky DJ, 
            Burmeister M
  TITLE     Mutation in ATG5 reduces autophagy and leads to ataxia with 
            developmental delay
  JOURNAL   Elife. 2016 Jan 26;5. pii: e12245. doi: 10.7554/eLife.12245.
  PUBMED    26812546
REFERENCE   2  (bases 1 to 7352)
  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: "Elife."; 
            date: "2016-01-26"; volume: "5. pii: e12245. doi"; pages: " 
            10.7554/eLife.12245"
COMMENT     Sequence Label: pLU-CMV-Flag-hATG5 E122D
FEATURES             Location/Qualifiers
     source          1..7352
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    111..227
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     enhancer        250..553
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        554..757
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     708..728
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     754..778
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      832..841
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             841..864
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     misc_feature    1723..2311
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1776..1796)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2194..2205)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             2398..2631
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    2703..2824
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2740..2759)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2794..2813
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      2864..2999
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(3031..3047)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3031..3047)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3044..3066)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3055..3071
                     /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(3079..3085,3086..3103,3104..3109))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3124..3145
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3262..3279)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3433..4021)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3513..3532)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4192..4983,4984..5052))
                     /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     4815..4834
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5053..5157)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5225..5243
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5281..5303)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5403..5422
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5616..5638
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5630..5647
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5631..5647
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        5662..5888
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             5889..6069
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    6116..6241
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6734..6967
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             7152..7196
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
ORIGIN
        1 ccaaccccga ggggacccga caggcccgaa ggaatagaag aagaaggtgg agagagagac
       61 agagacagat ccattcgatt agtgaacgga tctcgacggt atcggttaac ttttaaaaga
      121 aaagggggga ttggggggta cagtgcaggg gaaagaatag tagacataat agcaacagac
      181 atacaaacta aagaattaca aaaacaaatt acaaaattca aaattttatc gataagcttg
      241 ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc
      301 ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca
      361 ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta
      421 tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta
      481 tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat
      541 cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga
      601 ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca
      661 aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg
      721 taggcgtgta cggtgggagg tctatataag cagagctcgt ttagtgaacc gtcagatcgc
      781 ctggagacgc catccacgct gttttgacct ccatagaaga caccgactct agccaccatg
      841 gactacaaag acgatgacga caagtctaga atgacagatg acaaagatgt gcttcgagat
      901 gtgtggtttg gacgaattcc aacttgtttc acgctatatc aggatgagat aactgaaagg
      961 gaagcagaac catactattt gcttttgcca agagtaagtt atttgacgtt ggtaactgac
     1021 aaagtgaaaa agcactttca gaaggttatg agacaagaag acattagtga gatatggttt
     1081 gaatatgaag gcacaccact gaaatggcat tatccaattg gtttgctatt tgatcttctt
     1141 gcatcaagtt cagctcttcc ttggaacatc acagtacatt ttaagagttt tccagaaaaa
     1201 gaccttctgc actgtccatc taaggatgca attgacgctc attttatgtc atgtatgaaa
     1261 gaagctgatg ctttaaaaca taaaagtcaa gtaatcaatg aaatgcagaa aaaagatcac
     1321 aagcaactct ggatgggatt gcaaaatgac agatttgacc agttttgggc catcaatcgg
     1381 aaactcatgg aatatcctgc agaagaaaat ggatttcgtt atatcccctt tagaatatat
     1441 cagacaacga ctgaaagacc tttcattcag aagctgtttc gtcctgtggc tgcagatgga
     1501 cagttgcaca cactaggaga tctcctcaaa gaagtttgtc cttctgctat tgatcctgaa
     1561 gatggggaaa aaaagaatca agtgatgatt catggaattg agccaatgtt ggaaacacct
     1621 ctgcagtggc tgagtgaaca tctgagctac ccggataatt ttcttcatat tagtatcatc
     1681 ccacagccaa cagattgact cgaggaattc ggatccgtcg acaatcaacc tctggattac
     1741 aaaatttgtg aaagattgac tggtattctt aactatgttg ctccttttac gctatgtgga
     1801 tacgctgctt taatgccttt gtatcatgct attgcttccc gtatggcttt cattttctcc
     1861 tccttgtata aatcctggtt gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa
     1921 cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca ctggttgggg cattgccacc
     1981 acctgtcagc tcctttccgg gactttcgct ttccccctcc ctattgccac ggcggaactc
     2041 atcgccgcct gccttgcccg ctgctggaca ggggctcggc tgttgggcac tgacaattcc
     2101 gtggtgttgt cggggaagct gacgtccttt ccatggctgc tcgcctgtgt tgccacctgg
     2161 attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc tcaatccagc ggaccttcct
     2221 tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc ttcgccttcg ccctcagacg
     2281 agtcggatct ccctttgggc cgcctccccg cctggaattc gagctcggta cctttaagac
     2341 caatgactta caaggcagct gtagatctta gccacttttt aaaagaaaag gggggactgg
     2401 aagggctaat tcactcccaa cgaagacaag atctgctttt tgcttgtact gggtctctct
     2461 ggttagacca gatctgagcc tgggagctct ctggctaact agggaaccca ctgcttaagc
     2521 ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg
     2581 gtaactagag atccctcaga cccttttagt cagtgtggaa aatctctagc agtagtagtt
     2641 catgtcatct tattattcag tatttataac ttgcaaagaa atgaatatca gagagtgaga
     2701 ggaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
     2761 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
     2821 cttatcatgt ctggctctag ctatcccgcc cctaactccg cccatcccgc ccctaactcc
     2881 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     2941 cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     3001 aggcttttgc gggcccaaat tcgtaatcat gtcatagctg tttcctgtgt gaaattgtta
     3061 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
     3121 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     3181 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     3241 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     3301 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     3361 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     3421 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     3481 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     3541 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     3601 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     3661 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     3721 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     3781 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     3841 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     3901 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     3961 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     4021 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     4081 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     4141 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     4201 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     4261 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     4321 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     4381 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     4441 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     4501 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     4561 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     4621 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     4681 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     4741 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     4801 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     4861 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     4921 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     4981 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     5041 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     5101 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     5161 atttccccga aaagtgccac ctgacgtcta agaaaccatt attatcatga cattaaccta
     5221 taaaaatagg cgtatcacga ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa
     5281 cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag
     5341 cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggct ggcttaacta
     5401 tgcggcatca gagcagattg tactgagagt gcaccatatg cggtgtgaaa taccgcacag
     5461 atgcgtaagg agaaaatacc gcatcaggcg ccattcgcca ttcaggctgc gcaactgttg
     5521 ggaagggcga tcggtgcggg cctcttcgct attacgccag ctggcgaaag ggggatgtgc
     5581 tgcaaggcga ttaagttggg taacgccagg gttttcccag tcacgacgtt gtaaaacgac
     5641 ggccagtgcc aagctgacgc gtgtagtctt atgcaatact cttgtagtct tgcaacatgg
     5701 taacgatgag ttagcaacat gccttacaag gagagaaaaa gcaccgtgca tgccgattgg
     5761 tggaagtaag gtggtacgat cgtgccttat taggaaggca acagacgggt ctgacatgga
     5821 ttggacgaac cactgaattg ccgcattgca gagatattgt atttaagtgc ctagctcgat
     5881 acataaacgg gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag
     5941 ggaacccact gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc
     6001 gtctgttgtg tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa
     6061 tctctagcag tggcgcccga acagggactt gaaagcgaaa gggaaaccag aggagctctc
     6121 tcgacgcagg actcggcttg ctgaagcgcg cacggcaaga ggcgaggggc ggcgactggt
     6181 gagtacgcca aaaattttga ctagcggagg ctagaaggag agagatgggt gcgagagcgt
     6241 cagtattaag cgggggagaa ttagatcgcg atgggaaaaa attcggttaa ggccaggggg
     6301 aaagaaaaaa tataaattaa aacatatagt atgggcaagc agggagctag aacgattcgc
     6361 agttaatcct ggcctgttag aaacatcaga aggctgtaga caaatactgg gacagctaca
     6421 accatccctt cagacaggat cagaagaact tagatcatta tataatacag tagcaaccct
     6481 ctattgtgtg catcaaagga tagagataaa agacaccaag gaagctttag acaagataga
     6541 ggaagagcaa aacaaaagta agaccaccgc acagcaagcg gccgctgatc ttcagacctg
     6601 gaggaggaga tatgagggac aattggagaa gtgaattata taaatataaa gtagtaaaaa
     6661 ttgaaccatt aggagtagca cccaccaagg caaagagaag agtggtgcag agagaaaaaa
     6721 gagcagtggg aataggagct ttgttccttg ggttcttggg agcagcagga agcactatgg
     6781 gcgcagcgtc aatgacgctg acggtacagg ccagacaatt attgtctggt atagtgcagc
     6841 agcagaacaa tttgctgagg gctattgagg cgcaacagca tctgttgcaa ctcacagtct
     6901 ggggcatcaa gcagctccag gcaagaatcc tggctgtgga aagataccta aaggatcaac
     6961 agctcctggg gatttggggt tgctctggaa aactcatttg caccactgct gtgccttgga
     7021 atgctagttg gagtaataaa tctctggaac agatttggaa tcacacgacc tggatggagt
     7081 gggacagaga aattaacaat tacacaagct taatacactc cttaattgaa gaatcgcaaa
     7141 accagcaaga aaagaatgaa caagaattat tggaattaga taaatgggca agtttgtgga
     7201 attggtttaa cataacaaat tggctgtggt atataaaatt attcataatg atagtaggag
     7261 gcttggtagg tttaagaata gtttttgctg tactttctat agtgaataga gttaggcagg
     7321 gatattcacc attatcgttt cagacccacc tc
//
