LOCUS       sequence-389450-        7044 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of human ATG3 H262A.
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_H262A
FEATURES             Location/Qualifiers
     source          1..7044
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        88..467
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        468..671
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     622..642
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          807..939
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     922..941
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     primer_bind     983..1002
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        983..1001
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1011..1020
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1017..1907
                     /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             1920..1940
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     primer_bind     complement(2945..2963)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    2995..3116
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3006..3025)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3060..3079
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3361..3718
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3361..3552
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3384..3404)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3569..3704
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3769..4563
                     /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(3823..3842)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4433..4452
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4627..4675
                     /note="synthetic polyadenylation signal"
     promoter        4831..4935
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4936..5004,5005..5796)
                     /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(5154..5173)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(5954..6542)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6034..6053)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    6658..6941
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to 
                     maintain plasmids as monomers"
ORIGIN
        1 atattggcta ttggccattg catacgttgt atctatatca taatatgtac atttatattg
       61 gctcatgtcc aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat
      121 caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg
      181 taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt
      241 atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac
      301 ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg
      361 acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact
      421 ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt
      481 ggcagtacac caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc
      541 ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc
      601 gtaataaccc cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata
      661 taagcagagc tggtttagtg aaccgtcaga tcactagaag ctttattgcg gtagtttatc
      721 acagttaaat tgctaacgca gtcagtgctt ctgacacaac agtctcgaac ttaagctgca
      781 gaagttggtc gtgaggcact gggcaggtaa gtatcaaggt tacaagacag gtttaaggag
      841 accaatagaa actgggcttg tcgagacaga gaagactctt gcgtttctga taggcaccta
      901 ttggtcttac tgacatccac tttgcctttc tctccacagg tgtccactcc cagttcaatt
      961 acagctctta aggctagagt attaatacga ctcactatag ggctagcaaa gccaccatgg
     1021 cagaaatcgg tactggcttt ccattcgacc cccattatgt ggaagtcctg ggcgagcgca
     1081 tgcactacgt cgatgttggt ccgcgcgatg gcacccctgt gctgttcctg cacggtaacc
     1141 cgacctcctc ctacgtgtgg cgcaacatca tcccgcatgt tgcaccgacc catcgctgca
     1201 ttgctccaga cctgatcggt atgggcaaat ccgacaaacc agacctgggt tatttcttcg
     1261 acgaccacgt ccgcttcatg gatgccttca tcgaagccct gggtctggaa gaggtcgtcc
     1321 tggtcattca cgactggggc tccgctctgg gtttccactg ggccaagcgc aatccagagc
     1381 gcgtcaaagg tattgcattt atggagttca tccgccctat cccgacctgg gacgaatggc
     1441 cagaatttgc ccgcgagacc ttccaggcct tccgcaccac cgacgtcggc cgcaagctga
     1501 tcatcgatca gaacgttttt atcgagggta cgctgccgat gggtgtcgtc cgcccgctga
     1561 ctgaagtcga gatggaccat taccgcgagc cgttcctgaa tcctgttgac cgcgagccac
     1621 tgtggcgctt cccaaacgag ctgccaatcg ccggtgagcc agcgaacatc gtcgcgctgg
     1681 tcgaagaata catggactgg ctgcaccagt cccctgtccc gaagctgctg ttctggggca
     1741 ccccaggcgt tctgatccca ccggccgaag ccgctcgcct ggccaaaagc ctgcctaact
     1801 gcaaggctgt ggacatcggc ccgggtctga atctgctgca agaagacaac ccggacctga
     1861 tcggcagcga gatcgcgcgc tggctgtcga cgctcgagat ttccggcgag ccaaccactg
     1921 aggatctgta ctttcagagc gataacgcga tcgcttccga attcggatcc atgcagaatg
     1981 tgattaatac tgtgaaggga aaggcactgg aagtggctga gtacctgacc ccggtcctca
     2041 aggaatcaaa gtttaaggaa acaggtgtaa ttaccccaga agagtttgtg gcagctggag
     2101 atcacctagt ccaccactgt ccaacatggc aatgggctac aggggaagaa ttgaaagtga
     2161 aggcatacct accaacaggc aaacaatttt tggtaaccaa aaatgtgccg tgctataagc
     2221 ggtgcaaaca gatggaatat tcagatgaat tggaagctat cattgaagaa gatgatggtg
     2281 atggcggatg ggtagataca tatcacaaca caggtattac aggaataacg gaagccgtta
     2341 aagagatcac actggaaaat aaggacaata taaggcttca agattgctca gcactatgtg
     2401 aagaggaaga agatgaagat gaaggagaag ctgcagatat ggaagaatat gaagagagtg
     2461 gattgttgga aacagatgag gctaccctag atacaaggaa aatagtagaa gcttgtaaag
     2521 ccaaaactga tgctggcggt gaagatgcta ttttgcaaac cagaacttat gacctttaca
     2581 tcacttatga taaatattac cagactccac gattatggtt gtttggctat gatgagcaac
     2641 ggcagccttt aacagttgag cacatgtatg aagacatcag tcaggatcat gtgaagaaaa
     2701 cagtgaccat tgaaaatcac cctcatctgc caccacctcc catgtgttca gttgcgccat
     2761 gcaggcatgc tgaggtgatg aagaaaatca ttgagactgt tgcagaagga gggggagaac
     2821 ttggagttca tatgtatctt cttattttct tgaaatttgt acaagctgtc attccaacaa
     2881 tagaatatga ctacacaaga cacttcacaa tgtaagcggc cgcttgctga gttggctgct
     2941 gccaccgctg agcaataact agcataaccc cttggccgct tcgagcagac atgataagat
     3001 acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg
     3061 aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca
     3121 acaacaattg cattcatttt atgtttcagg ttcaggggga gatgtgggag gtttttttaa
     3181 gcaagtaaaa cctctacaaa tgtggtaaaa tcgaatttta acaaaatatt aacgcttaca
     3241 atttcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatacgcg
     3301 gatctgcgca gcaccatggc ctgaaataac ctctgaaaga ggaacttggt taggtacctt
     3361 ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
     3421 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
     3481 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     3541 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     3601 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc
     3661 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
     3721 gcttgattct tctgacacaa cagtctcgaa cttaaggcta gagccaccat gattgaacaa
     3781 gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg
     3841 gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc
     3901 ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca
     3961 gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc
     4021 actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca
     4081 tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat
     4141 acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca
     4201 cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg
     4261 ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc
     4321 gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct
     4381 ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct
     4441 acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac
     4501 ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc
     4561 tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgat
     4621 ggccgcaata aaatatcttt attttcatta catctgtgtg ttggtttttt gtgtgaatcg
     4681 atagcgataa ggatcctctt tgcgcttgcg ttttcccttg tccagatagc ccagtagctg
     4741 acattcatcc ggggtcagca ccgtttctgc ggactggctt tctacgtaat ggtttcttag
     4801 acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa
     4861 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat
     4921 tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg
     4981 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa
     5041 gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt
     5101 gagagttttc gccccgaaga acgttttcca atgatgagca ctttcaaagt tctgctatgt
     5161 ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat
     5221 tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg
     5281 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta
     5341 cttctgacaa ctatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat
     5401 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag
     5461 cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa
     5521 ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca
     5581 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc
     5641 ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt
     5701 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc
     5761 gctgagatag gtgcctcact gattaagcat tggtaattcg aaatgaccga ccaagcgacg
     5821 cccaaccggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     5881 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     5941 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     6001 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     6061 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     6121 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     6181 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     6241 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     6301 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     6361 tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc
     6421 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     6481 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatttc
     6541 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     6601 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttatagtccg
     6661 gaaatacagg aacgcacgct ggatggccct tcgctgggat ggtgaaacca tgaaaaatgg
     6721 cagcttcagt ggattaagtg ggggtaatgt ggcctgtacc ctctggttgc ataggtattc
     6781 atacggttaa aatttatcag gcgcgattgc ggcagttttt cgggtggttt gttgccattt
     6841 ttacctgtct gctgccgtga tcgcgctgaa cgcgttttag cggtgcgtac aattaaggga
     6901 ttatggtaaa tccacttact gtctgccctc gtagccatcg agataaaccg cagtactccg
     6961 gccacgatgc gtccggcgta gaggatcgag atcttcaata ttggccatta gccatattat
     7021 tcattggtta tatagcataa atca
//
