LOCUS       sequence-389405-        7044 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Mammalian Expression of human ATG3 H266Q.
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 Apr 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_H266Q
FEATURES             Location/Qualifiers
     source          1..7044
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        115..494
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        495..698
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     649..669
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          834..966
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     949..968
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     primer_bind     1010..1029
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1010..1028
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1038..1047
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1044..1934
                     /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             1947..1967
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     primer_bind     complement(2972..2990)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    3022..3143
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3033..3052)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3087..3106
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3388..3745
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3388..3579
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3411..3431)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3596..3731
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3796..4590
                     /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(3850..3869)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4460..4479
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4654..4702
                     /note="synthetic polyadenylation signal"
     promoter        4858..4962
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4963..5031,5032..5823)
                     /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(5181..5200)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(5981..6569)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6061..6080)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    6685..6968
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to 
                     maintain plasmids as monomers"
ORIGIN
        1 tattcattgg ttatatagca taaatcaata ttggctattg gccattgcat acgttgtatc
       61 tatatcataa tatgtacatt tatattggct catgtccaat atgaccgcca tgttggcatt
      121 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      181 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      241 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      301 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      361 atcatatgcc aagtccgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      421 atgcccagta catgacctta cgggactttc ctacttggca gtacatctac gtattagtca
      481 tcgctattac catggtgatg cggttttggc agtacaccaa tgggcgtgga tagcggtttg
      541 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      601 aaaatcaacg ggactttcca aaatgtcgta ataaccccgc cccgttgacg caaatgggcg
      661 gtaggcgtgt acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatca
      721 ctagaagctt tattgcggta gtttatcaca gttaaattgc taacgcagtc agtgcttctg
      781 acacaacagt ctcgaactta agctgcagaa gttggtcgtg aggcactggg caggtaagta
      841 tcaaggttac aagacaggtt taaggagacc aatagaaact gggcttgtcg agacagagaa
      901 gactcttgcg tttctgatag gcacctattg gtcttactga catccacttt gcctttctct
      961 ccacaggtgt ccactcccag ttcaattaca gctcttaagg ctagagtatt aatacgactc
     1021 actatagggc tagcaaagcc accatggcag aaatcggtac tggctttcca ttcgaccccc
     1081 attatgtgga agtcctgggc gagcgcatgc actacgtcga tgttggtccg cgcgatggca
     1141 cccctgtgct gttcctgcac ggtaacccga cctcctccta cgtgtggcgc aacatcatcc
     1201 cgcatgttgc accgacccat cgctgcattg ctccagacct gatcggtatg ggcaaatccg
     1261 acaaaccaga cctgggttat ttcttcgacg accacgtccg cttcatggat gccttcatcg
     1321 aagccctggg tctggaagag gtcgtcctgg tcattcacga ctggggctcc gctctgggtt
     1381 tccactgggc caagcgcaat ccagagcgcg tcaaaggtat tgcatttatg gagttcatcc
     1441 gccctatccc gacctgggac gaatggccag aatttgcccg cgagaccttc caggccttcc
     1501 gcaccaccga cgtcggccgc aagctgatca tcgatcagaa cgtttttatc gagggtacgc
     1561 tgccgatggg tgtcgtccgc ccgctgactg aagtcgagat ggaccattac cgcgagccgt
     1621 tcctgaatcc tgttgaccgc gagccactgt ggcgcttccc aaacgagctg ccaatcgccg
     1681 gtgagccagc gaacatcgtc gcgctggtcg aagaatacat ggactggctg caccagtccc
     1741 ctgtcccgaa gctgctgttc tggggcaccc caggcgttct gatcccaccg gccgaagccg
     1801 ctcgcctggc caaaagcctg cctaactgca aggctgtgga catcggcccg ggtctgaatc
     1861 tgctgcaaga agacaacccg gacctgatcg gcagcgagat cgcgcgctgg ctgtcgacgc
     1921 tcgagatttc cggcgagcca accactgagg atctgtactt tcagagcgat aacgcgatcg
     1981 cttccgaatt cggatccatg cagaatgtga ttaatactgt gaagggaaag gcactggaag
     2041 tggctgagta cctgaccccg gtcctcaagg aatcaaagtt taaggaaaca ggtgtaatta
     2101 ccccagaaga gtttgtggca gctggagatc acctagtcca ccactgtcca acatggcaat
     2161 gggctacagg ggaagaattg aaagtgaagg catacctacc aacaggcaaa caatttttgg
     2221 taaccaaaaa tgtgccgtgc tataagcggt gcaaacagat ggaatattca gatgaattgg
     2281 aagctatcat tgaagaagat gatggtgatg gcggatgggt agatacatat cacaacacag
     2341 gtattacagg aataacggaa gccgttaaag agatcacact ggaaaataag gacaatataa
     2401 ggcttcaaga ttgctcagca ctatgtgaag aggaagaaga tgaagatgaa ggagaagctg
     2461 cagatatgga agaatatgaa gagagtggat tgttggaaac agatgaggct accctagata
     2521 caaggaaaat agtagaagct tgtaaagcca aaactgatgc tggcggtgaa gatgctattt
     2581 tgcaaaccag aacttatgac ctttacatca cttatgataa atattaccag actccacgat
     2641 tatggttgtt tggctatgat gagcaacggc agcctttaac agttgagcac atgtatgaag
     2701 acatcagtca ggatcatgtg aagaaaacag tgaccattga aaatcaccct catctgccac
     2761 cacctcccat gtgttcagtt cacccatgca ggcaagctga ggtgatgaag aaaatcattg
     2821 agactgttgc agaaggaggg ggagaacttg gagttcatat gtatcttctt attttcttga
     2881 aatttgtaca agctgtcatt ccaacaatag aatatgacta cacaagacac ttcacaatgt
     2941 aagcggccgc ttgctgagtt ggctgctgcc accgctgagc aataactagc ataacccctt
     3001 ggccgcttcg agcagacatg ataagataca ttgatgagtt tggacaaacc acaactagaa
     3061 tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca
     3121 ttataagctg caataaacaa gttaacaaca acaattgcat tcattttatg tttcaggttc
     3181 agggggagat gtgggaggtt tttttaagca agtaaaacct ctacaaatgt ggtaaaatcg
     3241 aattttaaca aaatattaac gcttacaatt tcctgatgcg gtattttctc cttacgcatc
     3301 tgtgcggtat ttcacaccgc atacgcggat ctgcgcagca ccatggcctg aaataacctc
     3361 tgaaagagga acttggttag gtaccttctg aggcggaaag aaccagctgt ggaatgtgtg
     3421 tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     3481 tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     3541 gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc
     3601 gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat
     3661 ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt
     3721 ttttggaggc ctaggctttt gcaaaaagct tgattcttct gacacaacag tctcgaactt
     3781 aaggctagag ccaccatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg
     3841 gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc
     3901 gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt
     3961 gccctgaatg aactgcagga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt
     4021 ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc
     4081 gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc
     4141 atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac
     4201 caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag
     4261 gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag
     4321 gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat
     4381 atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg
     4441 gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa
     4501 tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc
     4561 ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgaa atgaccgacc
     4621 aagcgacgcc caacctgcca tcacgatggc cgcaataaaa tatctttatt ttcattacat
     4681 ctgtgtgttg gttttttgtg tgaatcgata gcgataagga tcctctttgc gcttgcgttt
     4741 tcccttgtcc agatagccca gtagctgaca ttcatccggg gtcagcaccg tttctgcgga
     4801 ctggctttct acgtaatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc
     4861 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     4921 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     4981 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     5041 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     5101 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     5161 atgagcactt tcaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     5221 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     5281 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     5341 atgagtgata acactgcggc caacttactt ctgacaacta tcggaggacc gaaggagcta
     5401 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     5461 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     5521 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     5581 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     5641 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     5701 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     5761 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     5821 taattcgaaa tgaccgacca agcgacgccc aaccggtatc agctcactca aaggcggtaa
     5881 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     5941 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     6001 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     6061 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     6121 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     6181 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     6241 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     6301 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     6361 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     6421 ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     6481 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     6541 agattacgcg cagaaaaaaa ggatttcaag aagatccttt gatcttttct acggggtctg
     6601 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     6661 tcttcaccta gatcctttta tagtccggaa atacaggaac gcacgctgga tggcccttcg
     6721 ctgggatggt gaaaccatga aaaatggcag cttcagtgga ttaagtgggg gtaatgtggc
     6781 ctgtaccctc tggttgcata ggtattcata cggttaaaat ttatcaggcg cgattgcggc
     6841 agtttttcgg gtggtttgtt gccattttta cctgtctgct gccgtgatcg cgctgaacgc
     6901 gttttagcgg tgcgtacaat taagggatta tggtaaatcc acttactgtc tgccctcgta
     6961 gccatcgaga taaaccgcag tactccggcc acgatgcgtc cggcgtagag gatcgagatc
     7021 ttcaatattg gccattagcc atat
//
