LOCUS       sequence-331416-        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        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     misc_feature    join(2076..2650,2651..2653,2654..2662)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2243..2260)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2470..2489
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(2663..2665,2666..2668,2669..3382)
                     /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(2663..2665,2666..2668,2669..3379)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2708..2729)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2969..2988)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3316..3337
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3505..3626
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3542..3561)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3596..3615
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3633..4088)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3770..3791)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3982..4001
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4115..4219
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4221..4578
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4221..4412
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4244..4264)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4429..4564
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4613..5407
                     /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             4613..5407
                     /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(4667..4686)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5277..5296
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5595..5614)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5639..5686
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6015..6603
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6504..6523
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctcgagct caagcttcga attcgtcgac tggatccggt accgaggaga
      661 tctgccgccg cgatcgccat gaggaaaagt ccaggtctgt ctgactgtct ttgggcctgg
      721 atcctccttc tgagcacact gactggaaga agctatggac agccgtcatt acaagatgaa
      781 cttaaagaca ataccactgt cttcaccagg attttggaca gactcctaga tggttatgac
      841 aatcgcctga gaccaggatt gggagagcgt gtaaccgaag tgaagactga tatcttcgtc
      901 accagtttcg gacccgtttc agaccatgat atggaatata caatagatgt atttttccgt
      961 caaagctgga aggatgaaag gttaaaattt aaaggaccta tgacagtcct ccggttaaat
     1021 aacctaatgg caagtaaaat ctggactccg gacacatttt tccacaatgg aaagaagtca
     1081 gtggcccaca acatgaccat gcccaacaaa ctcctgcgga tcacagagga tggcaccttg
     1141 ctgtacacca tgaggctgac agtgagagct gaatgtccga tgcatttgga ggacttccct
     1201 atggatgccc atgcttgccc actaaaattt ggaagttatg cttatacaag agcagaagtt
     1261 gtttatgaat ggaccagaga gccagcacgc tcagtggttg tagcagaaga tggatcacgt
     1321 ctaaaccagt atgaccttct tggacaaaca gtagactctg gaattgtcca gtcaagtaca
     1381 ggagaatatg ttgttatgac cactcatttc cacttgaaga gaaagattgg ctactttgtt
     1441 attcaaacat acctgccatg cataatgaca gtgattctct cacaagtctc cttctggctc
     1501 aacagagagt ctgtaccagc aagaactgtc tttggagtaa caactgtgct caccatgaca
     1561 acattgagca tcagtgccag aaactccctc cctaaggtgg cttatgcaac agctatggat
     1621 tggtttattg ccgtgtgcta tgactttgtg ttctcagctc tgattgagtt tgccacagta
     1681 aactatttca ctaagagagg ttatgcatgg gatggcaaaa gtgtggttcc agaaaagcca
     1741 aagaaagtaa aggatcctct tattaagaaa aacaacactt acgctccaac agcaaccagc
     1801 tacaccccta atttggccag gggcgacccg ggcttagcca ccattgctaa aagtgcaacc
     1861 atagaaccta aagaggtcaa gcccgaaaca aaaccaccag aacccaagaa aacctttaac
     1921 agtgtcagca aaattgaccg actgtcaaga atagccttcc cgctgctatt tggaatcttt
     1981 aacttagtct actgggctac gtatttaaac agagagcctc agctaaaagc ccccacacca
     2041 catcaataaa cggccggccg cgggcccggg atccgcccct ctccctcccc cccccctaac
     2101 gttactggcc gaagccgctt ggaataaggc cggtgtgcgt ttgtctatat gttattttcc
     2161 accatattgc cgtcttttgg caatgtgagg gcccggaaac ctggccctgt cttcttgacg
     2221 agcattccta ggggtctttc ccctctcgcc aaaggaatgc aaggtctgtt gaatgtcgtg
     2281 aaggaagcag ttcctctgga agcttcttga agacaaacaa cgtctgtagc gaccctttgc
     2341 aggcagcgga accccccacc tggcgacagg tgcctctgcg gccaaaagcc acgtgtataa
     2401 gatacacctg caaaggcggc acaaccccag tgccacgttg tgagttggat agttgtggaa
     2461 agagtcaaat ggctctcctc aagcgtattc aacaaggggc tgaaggatgc ccagaaggta
     2521 ccccattgta tgggatctga tctggggcct cggtgcacat gctttacatg tgtttagtcg
     2581 aggttaaaaa aacgtctagg ccccccgaac cacggggacg tggttttcct ttgaaaaaca
     2641 cgatgataat atggccacaa ccatggtgag caagggcgag gagctgttca ccggggtggt
     2701 gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga
     2761 gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa
     2821 gctgcccgtg ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag
     2881 ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta
     2941 cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt
     3001 gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga
     3061 ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat
     3121 catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga
     3181 ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc
     3241 cgtgctgctg cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa
     3301 cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg
     3361 catggacgag ctgtacaagt aaagcggccg cgactctaga tcataatcag ccataccaca
     3421 tttgtagagg ttttacttgc tttaaaaaac ctcccacacc tccccctgaa cctgaaacat
     3481 aaaatgaatg caattgttgt tgttaacttg tttattgcag cttataatgg ttacaaataa
     3541 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     3601 ttgtccaaac tcatcaatgt atcttaaggc gtaaattgta agcgttaata ttttgttaaa
     3661 attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa
     3721 aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa
     3781 caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca
     3841 gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg
     3901 taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc
     3961 ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc
     4021 aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca
     4081 gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     4141 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     4201 atattgaaaa aggaagagtc ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta
     4261 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     4321 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     4381 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
     4441 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     4501 gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     4561 ggcctaggct tttgcaaaga tcgatcaaga gacaggatga ggatcgtttc gcatgattga
     4621 acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga
     4681 ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg
     4741 gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaagacga
     4801 ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt
     4861 tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct
     4921 gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct
     4981 gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg
     5041 agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca
     5101 ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg agcatgcccg acggcgagga
     5161 tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt
     5221 ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt
     5281 ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct
     5341 ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt
     5401 cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca
     5461 cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg
     5521 gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccaccct
     5581 agggggaggc taactgaaac acggaaggag acaataccgg aaggaacccg cgctatgacg
     5641 gcaataaaaa gacagaataa aacgcacggt gttgggtcgt ttgttcataa acgcggggtt
     5701 cggtcccagg gctggcactc tgtcgatacc ccaccgagac cccattgggg ccaatacgcc
     5761 cgcgtttctt ccttttcccc accccacccc ccaagttcgg gtgaaggccc agggctcgca
     5821 gccaacgtcg gggcggcagg ccctgccata gcctcaggtt actcatatat actttagatt
     5881 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     5941 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     6001 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     6061 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     6121 aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag
     6181 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     6241 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     6301 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     6361 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     6421 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     6481 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     6541 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     6601 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     6661 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc catgcat
//
