LOCUS       sequence-332049-        6347 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Human ATG3.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6347)
  AUTHORS   Fracchiolla D, Chang C, Hurley JH, Martens S
  TITLE     A PI3K-WIPI2 positive feedback loop allosterically activates LC3 
            lipidation in autophagy.
  JOURNAL   J Cell Biol. 2020 Jul 6;219(7). pii: 151802. doi: 
            10.1083/jcb.201912098.
  PUBMED    32437499
REFERENCE   2  (bases 1 to 6347)
  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: "J Cell 
            Biol."; date: "2020-07-6"; volume: "219(7). pii: 151802. doi"; 
            pages: " 10.1083/jcb.201912098"
COMMENT     Sequence Label: 6xHis-TEV-ATG3 (Human autophagy E2-like enzyme)
FEATURES             Location/Qualifiers
     source          1..6347
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     969..988
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1008..1085)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1291..1310
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1398..1416
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1417..1441
                     /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)."
     RBS             1456..1478
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1497..1514
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1527..1547
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     promoter        2558..2576
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    2577..2601
                     /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)."
     CDS             2710..2754
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(2792..2810)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2806..2853
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2890..3345
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2977..2996)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3187..3208
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(join(3460..4251,4252..4320))
                     /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     4083..4102
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4321..4413)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4494..5082
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4983..5002
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5236..5253
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5268..5407
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5333..5352)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5493..5515
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5509..5700)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(6272..6347,1..1007))
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
ORIGIN
        1 gtttgcgtat tgggcgccag ggtggttttt cttttcacca gtgagacggg caacagctga
       61 ttgcccttca ccgcctggcc ctgagagagt tgcagcaagc ggtccacgct ggtttgcccc
      121 agcaggcgaa aatcctgttt gatggtggtt aacggcggga tataacatga gctgtcttcg
      181 gtatcgtcgt atcccactac cgagatgtcc gcaccaacgc gcagcccgga ctcggtaatg
      241 gcgcgcattg cgcccagcgc catctgatcg ttggcaacca gcatcgcagt gggaacgatg
      301 ccctcattca gcatttgcat ggtttgttga aaaccggaca tggcactcca gtcgccttcc
      361 cgttccgcta tcggctgaat ttgattgcga gtgagatatt tatgccagcc agccagacgc
      421 agacgcgccg agacagaact taatgggccc gctaacagcg cgatttgctg gtgacccaat
      481 gcgaccagat gctccacgcc cagtcgcgta ccgtcttcat gggagaaaat aatactgttg
      541 atgggtgtct ggtcagagac atcaagaaat aacgccggaa cattagtgca ggcagcttcc
      601 acagcaatgg catcctggtc atccagcgga tagttaatga tcagcccact gacgcgttgc
      661 gcgagaagat tgtgcaccgc cgctttacag gcttcgacgc cgcttcgttc taccatcgac
      721 accaccacgc tggcacccag ttgatcggcg cgagatttaa tcgccgcgac aatttgcgac
      781 ggcgcgtgca gggccagact ggaggtggca acgccaatca gcaacgactg tttgcccgcc
      841 agttgttgtg ccacgcggtt gggaatgtaa ttcagctccg ccatcgccgc ttccactttt
      901 tcccgcgttt tcgcagaaac gtggctggcc tggttcacca cgcgggaaac ggtctgataa
      961 gagacaccgg catactctgc gacatcgtat aacgttactg gtttcacatt caccaccctg
     1021 aattgactct cttccgggcg ctatcatgcc ataccgcgaa aggttttgcg ccattcgatg
     1081 gtgtccggga tctcgacgct ctcccttatg cgactcctgc attaggaagc agcccagtag
     1141 taggttgagg ccgttgagca ccgccgccgc aaggaatggt gcatgcaagg agatggcgcc
     1201 caacagtccc ccggccacgg ggcctgccac catacccacg ccgaaacaag cgctcatgag
     1261 cccgaagtgg cgagcccgat cttccccatc ggtgatgtcg gcgatatagg cgccagcaac
     1321 cgcacctgtg gcgccggtga tgccggccac gatgcgtccg gcgtagagga tcgagatcga
     1381 tctcgatccc gcgaaattaa tacgactcac tataggggaa ttgtgagcgg ataacaattc
     1441 ccctctagaa ataattttgt ttaactttaa gaaggagata taccatgggc agcagccatc
     1501 accatcatca ccacagccag gatccggaaa acctgtattt tcagggaatg cagaatgtga
     1561 ttaatactgt gaagggaaag gcactggaag tggctgagta cctgaccccg gtcctcaagg
     1621 aatcaaagtt taaggaaaca ggtgtaatta ccccagaaga gtttgtggca gctggagatc
     1681 acctagtcca ccactgtcca acatggcaat gggctacagg ggaagaattg aaagtgaagg
     1741 catacctacc aacaggcaaa caatttttgg taaccaaaaa tgtgccgtgc tataagcggt
     1801 gcaaacagat ggaatattca gatgaattgg aagctatcat tgaagaagat gatggtgatg
     1861 gcggatgggt agatacatat cacaacacag gtattacagg aataacggaa gccgttaaag
     1921 agatcacact ggaaaataag gacaatataa ggcttcaaga ttgctcagca ctatgtgaag
     1981 aggaagaaga tgaagatgaa ggagaagctg cagatatgga agaatatgaa gagagtggat
     2041 tgttggaaac agatgaggct accctagata caaggaaaat agtagaagct tgtaaagcca
     2101 aaactgatgc tggcggtgaa gatgctattt tgcaaaccag aacttatgac ctttacatca
     2161 cttatgataa atattaccag actccacgat tatggttgtt tggctatgat gagcaacggc
     2221 agcctttaac agttgagcac atgtatgaag acatcagtca ggatcatgtg aagaaaacag
     2281 tgaccattga aaatcaccct catctgccac cacctcccat gtgttcagtt cacccatgca
     2341 ggcatgctga ggtgatgaag aaaatcattg agactgttgc agaaggaggg ggagaacttg
     2401 gagttcatat gtatcttctt attttcttga aatttgtaca agctgtcatt ccaacaatag
     2461 aatatgacta cacaagacac ttcacaatgt aagcggccgc ataatgctta agtcgaacag
     2521 aaagtaatcg tattgtacac ggccgcataa tcgaaattaa tacgactcac tataggggaa
     2581 ttgtgagcgg ataacaattc cccatcttag tatattagtt aagtataaga aggagatata
     2641 catatggcag atctcaattg gatatcggcc ggccacgcga tcgctgacgt cggtaccctc
     2701 gagtctggta aagaaaccgc tgctgcgaaa tttgaacgcc agcacatgga ctcgtctact
     2761 agcgcagctt aattaaccta ggctgctgcc accgctgagc aataactagc ataacccctt
     2821 ggggcctcta aacgggtctt gaggggtttt ttgctgaaag gaggaactat atccggattg
     2881 gcgaatggga cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca
     2941 gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct
     3001 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt
     3061 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac
     3121 gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct
     3181 ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt
     3241 ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac
     3301 aaaaatttaa cgcgaatttt aacaaaatat taacgtttac aatttctggc ggcacgatgg
     3361 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     3421 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     3481 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     3541 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     3601 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     3661 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     3721 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     3781 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     3841 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     3901 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     3961 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     4021 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     4081 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     4141 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     4201 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     4261 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     4321 actcttcctt tttcaatcat gattgaagca tttatcaggg ttattgtctc atgagcggat
     4381 acatatttga atgtatttag aaaaataaac aaataggtca tgaccaaaat cccttaacgt
     4441 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     4501 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     4561 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     4621 gcgcagatac caaatactgt ccttctagtg tagccgtagt taggccacca cttcaagaac
     4681 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     4741 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     4801 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     4861 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     4921 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     4981 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     5041 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     5101 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     5161 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
     5221 cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgcct gatgcggtat
     5281 tttctcctta cgcatctgtg cggtatttca caccgcaatg gtgcactctc agtacaatct
     5341 gctctgatgc cgcatagtta agccagtata cactccgcta tcgctacgtg actgggtcat
     5401 ggctgcgccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc
     5461 ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc
     5521 accgtcatca ccgaaacgcg cgaggcagct gcggtaaagc tcatcagcgt ggtcgtgaag
     5581 cgattcacag atgtctgcct gttcatccgc gtccagctcg ttgagtttct ccagaagcgt
     5641 taatgtctgg cttctgataa agcgggccat gttaagggcg gttttttcct gtttggtcac
     5701 tgatgcctcc gtgtaagggg gatttctgtt catgggggta atgataccga tgaaacgaga
     5761 gaggatgctc acgatacggg ttactgatga tgaacatgcc cggttactgg aacgttgtga
     5821 gggtaaacaa ctggcggtat ggatgcggcg ggaccagaga aaaatcactc agggtcaatg
     5881 ccagcgcttc gttaatacag atgtaggtgt tccacagggt agccagcagc atcctgcgat
     5941 gcagatccgg aacataatgg tgcagggcgc tgacttccgc gtttccagac tttacgaaac
     6001 acggaaaccg aagaccattc atgttgttgc tcaggtcgca gacgttttgc agcagcagtc
     6061 gcttcacgtt cgctcgcgta tcggtgattc attctgctaa ccagtaaggc aaccccgcca
     6121 gcctagccgg gtcctcaacg acaggagcac gatcatgcta gtcatgcccc gcgcccaccg
     6181 gaaggagctg actgggttga aggctctcaa gggcatcggt cgagatcccg gtgcctaatg
     6241 agtgagctaa cttacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     6301 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcg
//
