LOCUS       SK0655(pCMV-FLAG        6950 bp    DNA     circular SYN 22-OCT-2025
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 6950)
  AUTHORS   Institute for Basic Science
  TITLE     Direct Submission
  JOURNAL   Exported Oct 22, 2025 from SnapGene 8.1.1
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6950
                     /mol_type="other DNA"
                     /organism="recombinant plasmid"
     promoter        1..591
                     /label=CMV
                     /note="color: #ffffff; direction: RIGHT"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
                     /note="color: #ffffff"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
                     /note="color: #ffffff; direction: RIGHT"
     primer_bind     591..653
                     /label=HA-CRY2(EA9,NheI)-F
                     /note="color: black; sequence: 
                     GACTGCTAGCGCCACCATGGGATACCCATACGACGTGCCTGACTACGCCCCACCGGTCA
                     TGAAGATGGAC; added: 2022-02-13"
     primer_bind     591..650
                     /label=FLAG-CRY2(EA9,NheI)-F
                     /note="color: black; sequence: 
                     GACTGCTAGCGCCACCATGGGAGACTACAAGGATGACGACGATAAGCCACCGGTCATGA
                     AGATG; added: 2022-02-13"
     CDS             join(603..608,609..632)
                     /codon_start=1
                     /label=FLAG tag
                     /note="This feature has 2 segments:
                        1: 603 .. 608 / #99ccff
                        2: 609 .. 632 / #99ccff"
                     /translation="MGDYKDDDDK"
     primer_bind     641..666
                     /label=CRY2(EA9,AgeI)-F
                     /note="color: black; sequence: 
                     GACTACCGGTCGCCACCATGAAGATGGACAAAAAGACCATCG; added: 
                     2021-08-12"
     CDS             642..2135
                     /codon_start=1
                     /label=CRY2(E281A)
                     /note="color: #33cccc"
                     /translation="MKMDKKTIVWFRRDLRIEDNPALAAAAHEGSVFPVFIWCPEEEGQ
                     FYPGRASRWWMKQSLAHLSQSLKALGSDLTLIKTHNTISAILDCIRVTGATKVVFNHLY
                     DPVSLVRDHTVKEKLVERGISVQSYNGDLLYEPWEIYCEKGKPFTSFNSYWKKCLDMSI
                     ESVMLPPPWRLMPITAAAEAIWACSIEELGLENEAEKPSNALLTRAWSPGWSNADKLLN
                     EFIEKQLIDYAKNSKKVVGNSTSLLSPYLHFGEISVRHVFQCARMKQIIWARDKNSEGA
                     ESADLFLRGIGLREYSRYICFNFPFTHEQSLLSHLRFFPWDADVDKFKAWRQGRTGYPL
                     VDAGMRELWATGWMHNRIRVIVSSFAVKFLLLPWKWGMKYFWDTLLDADLECDILGWQY
                     ISGSIPDGHELDRLDNPALQGAKYDPEGEYIRQWLPELARLPTEWIHHPWDAPLTVLKA
                     SGVELGTNYAKPIVDIDTARELLAKAISRTREAQIMIGAA"
     primer_bind     complement(1459..1490)
                     /label=Fragment 1.REV
                     /note="color: black; sequence: 
                     GCTTTCGGCGCCTTCGCTGTTTTTATCGCGAG"
     primer_bind     complement(1459..1490)
                     /label=STIM1(E281A)-R
                     /note="color: black; sequence: 
                     GCTTTCGGCGCCTTCGCTGTTTTTATCGCGAG"
     primer_bind     1476..1505
                     /label=Fragment 2.FOR
                     /note="color: black; sequence: 
                     GAAGGCGCCGAAAGCGCCGATCTGTTCCTG"
     primer_bind     1476..1505
                     /label=PHR(E281A)-F
                     /note="color: black; sequence: 
                     GAAGGCGCCGAAAGCGCCGATCTGTTCCTG"
     primer_bind     complement(1651..1669)
                     /label=HA-CRY2(EA9)-R
                     /note="color: black; sequence: GCATCCACGAGTGGGTACC; added: 
                     2022-02-13"
     primer_bind     complement(1717..1742)
                     /label=CRY2(internal)-R
                     /note="color: black; sequence: GACAGCAAATGAGGACACGATTACCC; 
                     added: 2022-02-21"
     CDS             2136..2162
                     /codon_start=1
                     /label=A9
                     /note="color: #ffffff"
                     /translation="ARDPPDLDN"
     CDS             2256..3599
                     /codon_start=1
                     /label=STIM1(238-685)
                     /note="color: #ff0000"
                     /translation="KEHMKKMMKDLEGLHRAEQSLHDLQERLHKAQEEHRTVEVEKVHL
                     EKKLRDEINLAKQEAQRLKELREGTENERSRQKYAEEELEQVREALRKAEKELESHSSW
                     YAPEALQKWLQLTHEVEVQYYNIKKQNAEKQLLVAKEGAEKIKKKRNTLFGTFHVAHSS
                     SLDDVDHKILTAKQALSEVTAALRERLHRWQQIEILCGFQIVNNPGIHSLVAALNIDPS
                     WMGSTRPNPAHFIMTDDVDDMDEEIVSPLSMQSPSLQSSVRQRLTEPQHGLGSQRDLTH
                     SDSESSLHMSDRQRVAPKPPQMSRAADEALNAMTSNGSHRLIEGVHPGSLVEKLPDSPA
                     LAKKALLALNHGLDKAHSLMELSPSAPPGGSPHLDSSRSHSPSSPDPDTPSPVGDSRAL
                     QASRNTRIPHLAGKKAVAEEDNGSIGEETDSSPGRKKFPLKIFKKPLKK"
     polyA_signal    3738..3859
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
                     /note="color: #a6acb3"
     rep_origin      complement(3866..4321)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
     promoter        4348..4452
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: RIGHT"
     promoter        4454..4811
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
                     /note="color: #ffffff; direction: RIGHT"
     rep_origin      4662..4797
                     /label=SV40 ori
                     /note="SV40 origin of replication"
                     /note="color: #ffff00"
     CDS             4846..5640
                     /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))"
                     /note="color: #ccffcc"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     polyA_signal    5872..5919
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
                     /note="color: #a6acb3"
     rep_origin      6248..6836
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
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 gctagcgcca
      601 ccatgggaga ctacaaggat gacgacgata agccaccggt catgaagatg gacaaaaaga
      661 ccatcgtctg gtttcggaga gatttgagaa tagaagataa tcccgcgctc gccgccgcgg
      721 cccacgaggg ttccgtcttc cccgttttca tttggtgtcc tgaagaagaa ggccagtttt
      781 atcccggaag ggcctctagg tggtggatga agcaaagtct ggcccatctt agccagtcac
      841 tgaaagcact gggcagtgat cttaccctga tcaagacaca caataccatc tctgccatcc
      901 tcgactgcat cagggtgacc ggcgcaacga aagtcgtgtt taaccacctg tacgatccag
      961 ttagtctggt gcgcgaccac actgtgaagg agaagctggt ggaacggggg atcagtgtgc
     1021 agagctacaa cggggacctt ctgtacgagc catgggagat ctattgcgag aaagggaaac
     1081 cgttcacctc cttcaacagt tactggaaga aatgtttgga tatgtcaata gagtccgtta
     1141 tgttgccccc tccctggaga ctgatgccga ttactgctgc tgcagaggcc atctgggcct
     1201 gctccatcga ggaactcggt ctggaaaatg aagcagaaaa gccaagcaat gcacttctca
     1261 ctagagcctg gagccccggc tggtctaatg ccgacaagct gcttaacgag ttcatcgaaa
     1321 aacaactgat tgactacgcg aagaactcca agaaagtggt aggtaactca actagcttgc
     1381 tctctccata tctccatttt ggcgagattt ctgtccgcca tgtatttcag tgcgctcgga
     1441 tgaaacagat tatctgggct cgcgataaaa acagcgaagg cgccgaaagc gccgatctgt
     1501 tcctgcgagg gatcggactt cgggaatact cccggtatat atgtttcaac tttccattca
     1561 cacacgagca gagtctgttg tcccacctca ggttcttccc ctgggacgcc gatgtcgaca
     1621 aattcaaggc atggagacag ggaaggacag ggtacccact cgtggatgct ggcatgagag
     1681 agctctgggc tacaggctgg atgcacaacc gcatccgggt aatcgtgtcc tcatttgctg
     1741 tcaagtttct gctcctgcct tggaaatggg gaatgaagta cttttgggat acccttctcg
     1801 acgccgactt ggagtgtgac attctgggat ggcaatatat tagcgggtca attcctgacg
     1861 gccatgagtt ggacaggttg gacaatccgg ccttgcaggg agctaagtat gatcccgaag
     1921 gagagtatat tcgacagtgg ctccccgagc tggcccgact tcctacggag tggattcacc
     1981 atccttggga cgcaccactg acagtgctca aggcaagcgg ggtggagctg ggcaccaatt
     2041 acgctaagcc tatagttgat atagatacag cacgcgagct gctggctaaa gcgatctctc
     2101 gcactcggga ggcgcagatt atgatcggtg ctgccgccag ggaccccccc gacctcgaca
     2161 acgaattcag tgggggaggc ggagggggtg gttcaggtgg tggtggtggt ggtggttcgg
     2221 gtggtggcgg agggggaggc tccggactca gatctaagga gcacatgaag aagatgatga
     2281 aggacttgga ggggttacac cgagctgagc agagtctgca tgaccttcag gaaaggctgc
     2341 acaaggccca ggaggagcac cgcacagtgg aggtggagaa ggtccatctg gaaaagaagc
     2401 tgcgcgatga gatcaacctt gctaagcagg aagcccagcg gctgaaggag ctgcgggagg
     2461 gtactgagaa tgagcggagc cgccaaaaat atgctgagga ggagttggag caggttcggg
     2521 aggccttgag gaaagcagag aaggagctag aatctcacag ctcatggtat gctccagagg
     2581 cccttcagaa gtggctgcag ctgacacatg aggtggaggt gcaatattac aacatcaaga
     2641 agcaaaatgc tgagaagcag ctgctggtgg ccaaggaggg ggctgagaag ataaaaaaga
     2701 agagaaacac actctttggc accttccacg tggcccacag ctcttccctg gatgatgtag
     2761 atcataaaat tctaacagct aagcaagcac tgagcgaggt gacagcagca ttgcgggagc
     2821 gcctgcaccg ctggcaacag atcgagatcc tctgtggctt ccagattgtc aacaaccctg
     2881 gcatccactc actggtggct gccctcaaca tagaccccag ctggatgggc agtacacgcc
     2941 ccaaccctgc tcacttcatc atgactgacg acgtggatga catggatgag gagattgtgt
     3001 ctcccttgtc catgcagtcc cctagcctgc agagcagtgt tcggcagcgc ctgacggagc
     3061 cacagcatgg cctgggatct cagagggatt tgacccattc cgattcggag tcctccctcc
     3121 acatgagtga ccgccagcgt gtggccccca aacctcctca gatgagccgt gctgcagacg
     3181 aggctctcaa tgccatgact tccaatggca gccaccggct gatcgagggg gtccacccag
     3241 ggtctctggt ggagaaactg cctgacagcc ctgccctggc caagaaggca ttactggcgc
     3301 tgaaccatgg gctggacaag gcccacagcc tgatggagct gagcccctca gccccacctg
     3361 gtggctctcc acatttggat tcttcccgtt ctcacagccc cagctcccca gacccagaca
     3421 caccatctcc agttggggac agccgagccc tgcaagccag ccgaaacaca cgcattcccc
     3481 acctggctgg caagaaggct gtggctgagg aggataatgg ctctattggc gaggaaacag
     3541 actccagccc aggccggaag aagtttcctc tcaaaatctt taagaagcct cttaagaagt
     3601 aggaattggg atccaccgga tctagataac tgatcataat cagccatacc acatttgtag
     3661 aggttttact tgctttaaaa aacctcccac acctccccct gaacctgaaa cataaaatga
     3721 atgcaattgt tgttgttaac ttgtttattg cagcttataa tggttacaaa taaagcaata
     3781 gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca
     3841 aactcatcaa tgtatcttaa cgcgtaaatt gtaagcgtta atattttgtt aaaattcgcg
     3901 ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct
     3961 tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt
     4021 ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat
     4081 ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca
     4141 ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac
     4201 gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta
     4261 gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg
     4321 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     4381 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     4441 aaaaggaaga gtcctgaggc ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg
     4501 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     4561 caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     4621 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     4681 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     4741 aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     4801 gcttttgcaa agatcgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat
     4861 ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca
     4921 caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg
     4981 gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcaaga cgaggcagcg
     5041 cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact
     5101 gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct
     5161 caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg
     5221 cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt
     5281 actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc
     5341 gcgccagccg aactgttcgc caggctcaag gcgagcatgc ccgacggcga ggatctcgtc
     5401 gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga
     5461 ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc
     5521 cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt
     5581 atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga
     5641 gcgggactct ggggttcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt
     5701 tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg
     5761 gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac cctaggggga
     5821 ggctaactga aacacggaag gagacaatac cggaaggaac ccgcgctatg acggcaataa
     5881 aaagacagaa taaaacgcac ggtgttgggt cgtttgttca taaacgcggg gttcggtccc
     5941 agggctggca ctctgtcgat accccaccga gaccccattg gggccaatac gcccgcgttt
     6001 cttccttttc cccaccccac cccccaagtt cgggtgaagg cccagggctc gcagccaacg
     6061 tcggggcggc aggccctgcc atagcctcag gttactcata tatactttag attgatttaa
     6121 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     6181 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     6241 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     6301 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     6361 ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg tagttaggcc
     6421 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     6481 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     6541 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     6601 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     6661 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     6721 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     6781 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     6841 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     6901 ttcctgcgtt atcccctgat tctgtggata accgtattac cgccatgcat
//
