LOCUS       sequence-220961-        5627 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  ORAI1 channel with C-terminal YFP, carrying the activating mutation 
            T184M from TAM, and the mutations L273D-L276D leading to a 
            deficiency in STIM1-mediated gating..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5627)
  AUTHORS   Bulla M, Gyimesi G, Kim JH, Bhardwaj R, Hediger MA, Frieden M, 
            Demaurex N
  TITLE     ORAI1 channel gating and selectivity is differentially altered by 
            natural mutations in the first or third transmembrane domain.
  JOURNAL   J Physiol. 2018 Nov 1. doi: 10.1113/JP277079.
  PUBMED    30382595
REFERENCE   2  (bases 1 to 5627)
  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: "J Physiol. 
            2018 Nov 1. doi: 10.1113/JP277079."
COMMENT     Sequence Label: ORAI1-YFP T184M-L273D-L276D
FEATURES             Location/Qualifiers
     source          1..5627
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        305..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      857..866
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      1507..1516
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1513..1515,1516..1518,1519..2232)
                     /codon_start=1
                     /product="enhanced YFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1558..1579)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1819..1838)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2166..2187
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2355..2476
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2392..2411)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2446..2465
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2483..2938)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2620..2641)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2832..2851
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2965..3069
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3071..3428
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3071..3262
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3094..3114)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3279..3414
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3463..4257
                     /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
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQLIAFYRLLDEFF"
     primer_bind     complement(3517..3536)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4127..4146
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4445..4464)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4489..4536
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4865..5453
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5354..5373
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      541 ccggactcag atctcgagat gcatccggag cccgccccgc ccccgagccg tagcagtccc
      601 gagcttcccc caagcggcgg cagcaccacc agcggcagcc gccggagccg ccgccgcagc
      661 ggggacgggg agcccccggg ggccccgcca ccgccgccgt ccgccgtcac ctacccggac
      721 tggatcggcc agagttactc cgaggtgatg agcctcaacg agcactccat gcaggcgctg
      781 tcctggcgca agctctactt gagccgcgcc aagcttaaag cctccagccg gacctcggct
      841 ctgctctccg gcttcgccat ggtggcaatg gtggaggtgc agctggacgc tgaccacgac
      901 tacccaccgg ggctgctcat cgccttcagt gcctgcacca cagtgctggt ggctgtgcac
      961 ctgtttgcgc tcatgatcag cacctgcatc ctgcccaaca tcgaggcggt gagcaacgtg
     1021 cacaatctca actcggtcaa ggagtccccc catgagcgca tgcaccgcca catcgagctg
     1081 gcctgggcct tctccaccgt catcggcatg ctgctcttcc tagctgaggt ggtgctgctc
     1141 tgctgggtca agttcttgcc cctcaagaag cagccaggcc agccaaggcc caccagcaag
     1201 ccccccgcca gtggcgcagc agccaacgtc agcaccagcg gcatcacccc gggccaggca
     1261 gctgccatcg cctcgaccac catcatggtg cccttcggcc tgatctttat cgtcttcgcc
     1321 gtccacttct accgctcact ggttagccat aagaccgacc gacagttcca ggaggacaac
     1381 gaggacgcgg agtttgcccg cttacaggac cagctggacc acagagggga ccaccccctg
     1441 acgcccggca gccactatgc ccgaattctg cagtcgacgg taccgcgggc ccgggatcca
     1501 ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
     1561 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
     1621 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
     1681 ccctggccca ccctcgtgac caccttcggc tacggcctgc agtgcttcgc ccgctacccc
     1741 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
     1801 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
     1861 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     1921 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
     1981 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     2041 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     2101 cccgacaacc actacctgag ctaccagtcc aagctgagca aagaccccaa cgagaagcgc
     2161 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     2221 ctgtacaagt aaagcggccg cgactctaga tcataatcag ccataccaca tttgtagagg
     2281 ttttacttgc tttaaaaaac ctcccacacc tccccctgaa cctgaaacat aaaatgaatg
     2341 caattgttgt tgttaacttg tttattgcag cttataatgg ttacaaataa agcaatagca
     2401 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac
     2461 tcatcaatgt atcttaaggc gtaaattgta agcgttaata ttttgttaaa attcgcgtta
     2521 aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat
     2581 aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca
     2641 ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc
     2701 ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta
     2761 aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg
     2821 gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg
     2881 gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtca
     2941 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     3001 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     3061 aggaagagtc ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa
     3121 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     3181 ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
     3241 attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca
     3301 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg
     3361 ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct
     3421 tttgcaaaga tcgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga
     3481 ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa
     3541 cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt
     3601 ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaagacga ggcagcgcgg
     3661 ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa
     3721 gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac
     3781 cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt
     3841 gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact
     3901 cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg
     3961 ccagccgaac tgttcgccag gctcaaggcg agcatgcccg acggcgagga tctcgtcgtg
     4021 acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc
     4081 atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt
     4141 gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc
     4201 gccgctcccg attcgcagct catcgccttc tatcgccttc ttgacgagtt cttctgagcg
     4261 ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg
     4321 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct
     4381 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccaccct agggggaggc
     4441 taactgaaac acggaaggag acaataccgg aaggaacccg cgctatgacg gcaataaaaa
     4501 gacagaataa aacgcacggt gttgggtcgt ttgttcataa acgcggggtt cggtcccagg
     4561 gctggcactc tgtcgatacc ccaccgagac cccattgggg ccaatacgcc cgcgtttctt
     4621 ccttttcccc accccacccc ccaagttcgg gtgaaggccc agggctcgca gccaacgtcg
     4681 gggcggcagg ccctgccata gcctcaggtt actcatatat actttagatt gatttaaaac
     4741 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     4801 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     4861 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     4921 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     4981 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     5041 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     5101 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     5161 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     5221 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     5281 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     5341 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     5401 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     5461 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     5521 ctgcgttatc ccctgattct gtggataacc gtattaccgc catgcattag ttattaatag
     5581 taatcaatta cggggtcatt agttcatagc ccatatatgg agttccg
//
