LOCUS       sequence-338273-        6717 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  human GABAA receptors alpha1 subunit carrying A322D mutation and 
            IRES2-EGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6717)
  AUTHORS   Fu YL, Han DY, Wang YJ, Di XJ, Yu HB, Mu TW
  TITLE     Remodeling the endoplasmic reticulum proteostasis network restores 
            proteostasis of pathogenic GABAA receptors.
  JOURNAL   PLoS One. 2018 Nov 27;13(11):e0207948. doi: 
            10.1371/journal.pone.0207948. eCollection 2018.
  PUBMED    30481215
REFERENCE   2  (bases 1 to 6717)
  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: "PLoS One.";
            date: "2018-11-27"; volume: "13(11):e0207948. doi"; pages: " 
            10.1371/journal.pone.0207948. eCollection 2018"
COMMENT     Sequence Label: pCMV-GABRA1(A322D)-IRES2-EGFP
FEATURES             Location/Qualifiers
     source          1..6717
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        2..305
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        306..509
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     460..480
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     misc_feature    join(2017..2591,2592..2594,2595..2603)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2184..2201)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2411..2430
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(2604..2606,2607..2609,2610..3323)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2604..2606,2607..2609,2610..3320)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2649..2670)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2910..2929)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3257..3278
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3446..3567
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3483..3502)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3537..3556
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3574..4029)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3711..3732)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3923..3942
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4056..4160
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4162..4519
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4162..4353
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4185..4205)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4370..4505
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4554..5348
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             4554..5348
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4608..4627)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5218..5237
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5536..5555)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5580..5627
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5956..6544
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6445..6464
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat
       61 tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc
      121 aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc
      181 caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt
      241 acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta
      301 ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt gactcacggg
      361 gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac
      421 gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc ggtaggcgtg
      481 tacggtggga ggtctatata agcagagctg gtttagtgaa ccgtcagatc cgctagcgct
      541 accggactca gatctcgagc tcaagcttcg aattcgtcga ctggatccgg taccgaggag
      601 atctgccgcc gcgatcgcca tgaggaaaag tccaggtctg tctgactgtc tttgggcctg
      661 gatcctcctt ctgagcacac tgactggaag aagctatgga cagccgtcat tacaagatga
      721 acttaaagac aataccactg tcttcaccag gattttggac agactcctag atggttatga
      781 caatcgcctg agaccaggat tgggagagcg tgtaaccgaa gtgaagactg atatcttcgt
      841 caccagtttc ggacccgttt cagaccatga tatggaatat acaatagatg tatttttccg
      901 tcaaagctgg aaggatgaaa ggttaaaatt taaaggacct atgacagtcc tccggttaaa
      961 taacctaatg gcaagtaaaa tctggactcc ggacacattt ttccacaatg gaaagaagtc
     1021 agtggcccac aacatgacca tgcccaacaa actcctgcgg atcacagagg atggcacctt
     1081 gctgtacacc atgaggctga cagtgagagc tgaatgtccg atgcatttgg aggacttccc
     1141 tatggatgcc catgcttgcc cactaaaatt tggaagttat gcttatacaa gagcagaagt
     1201 tgtttatgaa tggaccagag agccagcacg ctcagtggtt gtagcagaag atggatcacg
     1261 tctaaaccag tatgaccttc ttggacaaac agtagactct ggaattgtcc agtcaagtac
     1321 aggagaatat gttgttatga ccactcattt ccacttgaag agaaagattg gctactttgt
     1381 tattcaaaca tacctgccat gcataatgac agtgattctc tcacaagtct ccttctggct
     1441 caacagagag tctgtaccag caagaactgt ctttggagta acaactgtgc tcaccatgac
     1501 aacattgagc atcagtgcca gaaactccct ccctaaggtg gcttatgcaa cagctatgga
     1561 ttggtttatt gccgtgtgct atgactttgt gttctcagct ctgattgagt ttgccacagt
     1621 aaactatttc actaagagag gttatgcatg ggatggcaaa agtgtggttc cagaaaagcc
     1681 aaagaaagta aaggatcctc ttattaagaa aaacaacact tacgctccaa cagcaaccag
     1741 ctacacccct aatttggcca ggggcgaccc gggcttagcc accattgcta aaagtgcaac
     1801 catagaacct aaagaggtca agcccgaaac aaaaccacca gaacccaaga aaacctttaa
     1861 cagtgtcagc aaaattgacc gactgtcaag aatagccttc ccgctgctat ttggaatctt
     1921 taacttagtc tactgggcta cgtatttaaa cagagagcct cagctaaaag cccccacacc
     1981 acatcaataa acggccggcc gcgggcccgg gatccgcccc tctccctccc ccccccctaa
     2041 cgttactggc cgaagccgct tggaataagg ccggtgtgcg tttgtctata tgttattttc
     2101 caccatattg ccgtcttttg gcaatgtgag ggcccggaaa cctggccctg tcttcttgac
     2161 gagcattcct aggggtcttt cccctctcgc caaaggaatg caaggtctgt tgaatgtcgt
     2221 gaaggaagca gttcctctgg aagcttcttg aagacaaaca acgtctgtag cgaccctttg
     2281 caggcagcgg aaccccccac ctggcgacag gtgcctctgc ggccaaaagc cacgtgtata
     2341 agatacacct gcaaaggcgg cacaacccca gtgccacgtt gtgagttgga tagttgtgga
     2401 aagagtcaaa tggctctcct caagcgtatt caacaagggg ctgaaggatg cccagaaggt
     2461 accccattgt atgggatctg atctggggcc tcggtacaca tgctttacat gtgtttagtc
     2521 gaggttaaaa aaacgtctag gccccccgaa ccacggggac gtggttttcc tttgaaaaac
     2581 acgatgataa tatggccaca accatggtga gcaagggcga ggagctgttc accggggtgg
     2641 tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg
     2701 agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca
     2761 agctgcccgt gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca
     2821 gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct
     2881 acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg
     2941 tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg
     3001 aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata
     3061 tcatggccga caagcagaag aacggcatca aggtgaactt caagatccgc cacaacatcg
     3121 aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc
     3181 ccgtgctgct gcccgacaac cactacctga gcacccagtc cgccctgagc aaagacccca
     3241 acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg
     3301 gcatggacga gctgtacaag taaagcggcc gcgactctag atcataatca gccataccac
     3361 atttgtagag gttttacttg ctttaaaaaa cctcccacac ctccccctga acctgaaaca
     3421 taaaatgaat gcaattgttg ttgttaactt gtttattgca gcttataatg gttacaaata
     3481 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     3541 tttgtccaaa ctcatcaatg tatcttaagg cgtaaattgt aagcgttaat attttgttaa
     3601 aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca
     3661 aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga
     3721 acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc
     3781 agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc
     3841 gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc
     3901 cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg
     3961 caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac
     4021 agggcgcgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     4081 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     4141 aatattgaaa aaggaagagt cctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt
     4201 agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     4261 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     4321 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct
     4381 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc
     4441 agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag gcttttttgg
     4501 aggcctaggc ttttgcaaag atcgatcaag agacaggatg aggatcgttt cgcatgattg
     4561 aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg
     4621 actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg
     4681 ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaagacg
     4741 aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg
     4801 ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc
     4861 tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc
     4921 tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc
     4981 gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc
     5041 aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gagcatgccc gacggcgagg
     5101 atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct
     5161 tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt
     5221 tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc
     5281 tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt
     5341 tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca acctgccatc
     5401 acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg
     5461 ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc
     5521 tagggggagg ctaactgaaa cacggaagga gacaataccg gaaggaaccc gcgctatgac
     5581 ggcaataaaa agacagaata aaacgcacgg tgttgggtcg tttgttcata aacgcggggt
     5641 tcggtcccag ggctggcact ctgtcgatac cccaccgaga ccccattggg gccaatacgc
     5701 ccgcgtttct tccttttccc caccccaccc cccaagttcg ggtgaaggcc cagggctcgc
     5761 agccaacgtc ggggcggcag gccctgccat agcctcaggt tactcatata tactttagat
     5821 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
     5881 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     5941 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     6001 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     6061 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
     6121 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     6181 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     6241 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     6301 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     6361 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     6421 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     6481 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     6541 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     6601 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg ccatgcatta
     6661 gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg gagttcc
//
