LOCUS       sequence-389123-        7044 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of human ATG3 H266A.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7044)
  TITLE     Hurley ATG3 plasmids
REFERENCE   2  (bases 1 to 7044)
  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"
COMMENT     Sequence Label: pHTN-Halo-TEV-ATG3_H266A
FEATURES             Location/Qualifiers
     source          1..7044
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    168..216
                     /note="synthetic polyadenylation signal"
     promoter        372..476
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(477..545,546..1337)
                     /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     complement(695..714)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(1495..2083)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1575..1594)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    2199..2482
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to 
                     maintain plasmids as monomers"
     enhancer        2673..3052
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3053..3256
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3207..3227
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          3392..3524
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     3507..3526
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     primer_bind     3568..3587
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3568..3586
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      3596..3605
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3602..4492
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="MAEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSS
                     YVWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVI
                     HDWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLII
                     DQNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALV
                     EEYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDL
                     IGSEIARWLSTLEISG"
     CDS             4505..4525
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     primer_bind     complement(5530..5548)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    5580..5701
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5591..5610)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5645..5664
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        5946..6303
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5946..6137
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5969..5989)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      6154..6289
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             join(6354..7044,1..104)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(6408..6427)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     7018..7037
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
ORIGIN
        1 agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt
       61 cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt
      121 cgaaatgacc gaccaagcga cgcccaacct gccatcacga tggccgcaat aaaatatctt
      181 tattttcatt acatctgtgt gttggttttt tgtgtgaatc gatagcgata aggatcctct
      241 ttgcgcttgc gttttccctt gtccagatag cccagtagct gacattcatc cggggtcagc
      301 accgtttctg cggactggct ttctacgtaa tggtttctta gacgtcaggt ggcacttttc
      361 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
      421 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
      481 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
      541 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
      601 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
      661 aacgttttcc aatgatgagc actttcaaag ttctgctatg tggcgcggta ttatcccgta
      721 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
      781 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
      841 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca actatcggag
      901 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
      961 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     1021 tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     1081 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     1141 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     1201 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     1261 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     1321 tgattaagca ttggtaattc gaaatgaccg accaagcgac gcccaaccgg tatcagctca
     1381 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     1441 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     1501 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     1561 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     1621 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     1681 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     1741 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     1801 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     1861 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     1921 cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     1981 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     2041 tgtttgcaag cagcagatta cgcgcagaaa aaaaggattt caagaagatc ctttgatctt
     2101 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     2161 attatcaaaa aggatcttca cctagatcct tttatagtcc ggaaatacag gaacgcacgc
     2221 tggatggccc ttcgctggga tggtgaaacc atgaaaaatg gcagcttcag tggattaagt
     2281 gggggtaatg tggcctgtac cctctggttg cataggtatt catacggtta aaatttatca
     2341 ggcgcgattg cggcagtttt tcgggtggtt tgttgccatt tttacctgtc tgctgccgtg
     2401 atcgcgctga acgcgtttta gcggtgcgta caattaaggg attatggtaa atccacttac
     2461 tgtctgccct cgtagccatc gagataaacc gcagtactcc ggccacgatg cgtccggcgt
     2521 agaggatcga gatcttcaat attggccatt agccatatta ttcattggtt atatagcata
     2581 aatcaatatt ggctattggc cattgcatac gttgtatcta tatcataata tgtacattta
     2641 tattggctca tgtccaatat gaccgccatg ttggcattga ttattgacta gttattaata
     2701 gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact
     2761 tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat
     2821 gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta
     2881 tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa gtccgccccc
     2941 tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttacg
     3001 ggactttcct acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg
     3061 gttttggcag tacaccaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct
     3121 ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa
     3181 atgtcgtaat aaccccgccc cgttgacgca aatgggcggt aggcgtgtac ggtgggaggt
     3241 ctatataagc agagctggtt tagtgaaccg tcagatcact agaagcttta ttgcggtagt
     3301 ttatcacagt taaattgcta acgcagtcag tgcttctgac acaacagtct cgaacttaag
     3361 ctgcagaagt tggtcgtgag gcactgggca ggtaagtatc aaggttacaa gacaggttta
     3421 aggagaccaa tagaaactgg gcttgtcgag acagagaaga ctcttgcgtt tctgataggc
     3481 acctattggt cttactgaca tccactttgc ctttctctcc acaggtgtcc actcccagtt
     3541 caattacagc tcttaaggct agagtattaa tacgactcac tatagggcta gcaaagccac
     3601 catggcagaa atcggtactg gctttccatt cgacccccat tatgtggaag tcctgggcga
     3661 gcgcatgcac tacgtcgatg ttggtccgcg cgatggcacc cctgtgctgt tcctgcacgg
     3721 taacccgacc tcctcctacg tgtggcgcaa catcatcccg catgttgcac cgacccatcg
     3781 ctgcattgct ccagacctga tcggtatggg caaatccgac aaaccagacc tgggttattt
     3841 cttcgacgac cacgtccgct tcatggatgc cttcatcgaa gccctgggtc tggaagaggt
     3901 cgtcctggtc attcacgact ggggctccgc tctgggtttc cactgggcca agcgcaatcc
     3961 agagcgcgtc aaaggtattg catttatgga gttcatccgc cctatcccga cctgggacga
     4021 atggccagaa tttgcccgcg agaccttcca ggccttccgc accaccgacg tcggccgcaa
     4081 gctgatcatc gatcagaacg tttttatcga gggtacgctg ccgatgggtg tcgtccgccc
     4141 gctgactgaa gtcgagatgg accattaccg cgagccgttc ctgaatcctg ttgaccgcga
     4201 gccactgtgg cgcttcccaa acgagctgcc aatcgccggt gagccagcga acatcgtcgc
     4261 gctggtcgaa gaatacatgg actggctgca ccagtcccct gtcccgaagc tgctgttctg
     4321 gggcacccca ggcgttctga tcccaccggc cgaagccgct cgcctggcca aaagcctgcc
     4381 taactgcaag gctgtggaca tcggcccggg tctgaatctg ctgcaagaag acaacccgga
     4441 cctgatcggc agcgagatcg cgcgctggct gtcgacgctc gagatttccg gcgagccaac
     4501 cactgaggat ctgtactttc agagcgataa cgcgatcgct tccgaattcg gatccatgca
     4561 gaatgtgatt aatactgtga agggaaaggc actggaagtg gctgagtacc tgaccccggt
     4621 cctcaaggaa tcaaagttta aggaaacagg tgtaattacc ccagaagagt ttgtggcagc
     4681 tggagatcac ctagtccacc actgtccaac atggcaatgg gctacagggg aagaattgaa
     4741 agtgaaggca tacctaccaa caggcaaaca atttttggta accaaaaatg tgccgtgcta
     4801 taagcggtgc aaacagatgg aatattcaga tgaattggaa gctatcattg aagaagatga
     4861 tggtgatggc ggatgggtag atacatatca caacacaggt attacaggaa taacggaagc
     4921 cgttaaagag atcacactgg aaaataagga caatataagg cttcaagatt gctcagcact
     4981 atgtgaagag gaagaagatg aagatgaagg agaagctgca gatatggaag aatatgaaga
     5041 gagtggattg ttggaaacag atgaggctac cctagataca aggaaaatag tagaagcttg
     5101 taaagccaaa actgatgctg gcggtgaaga tgctattttg caaaccagaa cttatgacct
     5161 ttacatcact tatgataaat attaccagac tccacgatta tggttgtttg gctatgatga
     5221 gcaacggcag cctttaacag ttgagcacat gtatgaagac atcagtcagg atcatgtgaa
     5281 gaaaacagtg accattgaaa atcaccctca tctgccacca cctcccatgt gttcagttca
     5341 cccatgcagg gcagctgagg tgatgaagaa aatcattgag actgttgcag aaggaggggg
     5401 agaacttgga gttcatatgt atcttcttat tttcttgaaa tttgtacaag ctgtcattcc
     5461 aacaatagaa tatgactaca caagacactt cacaatgtaa gcggccgctt gctgagttgg
     5521 ctgctgccac cgctgagcaa taactagcat aaccccttgg ccgcttcgag cagacatgat
     5581 aagatacatt gatgagtttg gacaaaccac aactagaatg cagtgaaaaa aatgctttat
     5641 ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt
     5701 taacaacaac aattgcattc attttatgtt tcaggttcag ggggagatgt gggaggtttt
     5761 tttaagcaag taaaacctct acaaatgtgg taaaatcgaa ttttaacaaa atattaacgc
     5821 ttacaatttc ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat
     5881 acgcggatct gcgcagcacc atggcctgaa ataacctctg aaagaggaac ttggttaggt
     5941 accttctgag gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg tggaaagtcc
     6001 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg
     6061 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     6121 tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
     6181 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
     6241 tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
     6301 aaaaagcttg attcttctga cacaacagtc tcgaacttaa ggctagagcc accatgattg
     6361 aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg
     6421 actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg
     6481 ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg
     6541 aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg
     6601 ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc
     6661 tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc
     6721 tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc
     6781 gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc
     6841 aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc gacggcgagg
     6901 atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct
     6961 tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt
     7021 tggctacccg tgatattgct gaag
//
