LOCUS       sequence-209272-        6570 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Genetically-encoded calcium actuator; mCherry-tagged LOV2-STIM1 
            chimera that enables optical control of calcium signals and calcium 
            dependent signaling.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6570)
  AUTHORS   He L, Zhang Y, Ma G, Tan P, Li Z, Zang S, Wu X, Jing J, Fang S, Zhou
            L, Wang Y, Huang Y, Hogan PG, Han G, Zhou Y
  TITLE     Near-infrared photoactivatable control of Ca(2+) signaling and 
            optogenetic immunomodulation.
  JOURNAL   Elife. 2015 Dec 8;4. pii: e10024. doi: 10.7554/eLife.10024.
  PUBMED    26646180
REFERENCE   2  (bases 1 to 6570)
  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: "Elife."; 
            date: "2015-12-8"; volume: "4. pii: e10024. doi"; pages: " 
            10.7554/eLife.10024"
COMMENT     Sequence Label: Opto-CRAC version 1
FEATURES             Location/Qualifiers
     source          1..6570
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_recomb     181..1008
                     /label=baculovirus recombination region (lef2/ORF603)
                     /note="contains ORF603 and part of lef2"
     CDS             181..305
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(343..948)
                     /codon_start=1
                     /product="baculovirus ORF603 protein"
                     /label=ORF603
                     /translation="MAVIFNNKQLLADNSIEKGGELFLFNGSYNILESYVNPVLLKNGV
                     IELEEAAYYAGNILYKTDDPKFIDYINLIIKATHSEELPENSTVVNYRKTMRSGTIHPI
                     KKDIYIYDNKKFTLYDRYIYGYDNNYVNFYEEKNEKEKEYEEEDDKASSLCENKIILSQ
                     INCESFENDFKYYLSDYNYAFSIIDNTTNVLVAFGLYR"
     enhancer        1076..1379
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1381..1583
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1534..1554
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1786..1808
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1870..1889
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     primer_bind     1931..1950
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1931..1949
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1950..1974
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        1990..2099
                     /label=p10 promoter
                     /note="baculovirus promoter for expression in insect cells"
     CDS             2120..2137
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2144..2848
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(2286..2304)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2554..2573
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             2855..3283
                     /codon_start=1
                     /gene="NPH1 from Avena sativa"
                     /product="light, oxygen, or voltage domain from phototropin
                     1 of Avena sativa (Herrou and Crosson, 2011)"
                     /label=LOV2
                     /note="photoswitch that undergoes a light-inducible 
                     conformational change"
                     /translation="LATTLERIEKNFVITDPRLPDNPIIFASDSFLQLTEYSREEILGR
                     NCRFLQGPETDRATVRKIRDAIDNQTEVTVQLINYTKSGKKFWNLFHLQPMRDQKGDVQ
                     YFIGVQLDGTEHVRDAAEREGVMLIKKTAENIDEAAKEL"
     CDS             3752..3775
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     primer_bind     complement(3865..3884)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3930..3985
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3984..4003)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     terminator      4073..4120
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     misc_recomb     4133..4838
                     /label=baculovirus recombination region (ORF1629)
                     /note="contains part of ORF1629"
     3'UTR           4221..4593
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     CDS             complement(4250..4838)
                     /codon_start=2
                     /product="baculovirus capsid-associated protein"
                     /label=ORF1629
                     /note="required for viral replication"
                     /translation="IPKSSTTNLIADVLADTINRRRVAMAKSSSEATSNDEGWDDDDNR
                     PNKANTPDVKYVQALFNVFTSSQLYTNDSDERNTKAHNILNDVEPLLQNKTQTNIDKAR
                     LLLQDLASFVALSENPLDSPAIGSEKQPLFETNRNLFYKSIEDLIFKFRYKDAENHLIF
                     ALTYHPKDYKFNELLKYVQQLSVNQQRTESSA"
     protein_bind    complement(4847..4880)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     rep_origin      complement(5048..5635)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5127..5146)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5648..6439,6440..6508))
                     /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     6271..6290
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 tcctgcatct tttaatcaaa tcccaagatg tgtataaacg cgccggtatg tacaggaaga
       61 ggtttatact aaactgttac attgcaaacg tggtttcgtg tgccaagtgt gaaaaccgat
      121 gtttaatcaa ggctctgacg catttctaca accacgactc caagtgtgtg ggtgaagtca
      181 tgcatctttt aatcaaatcc caagatgtgt ataaaccacc aaactgccaa aaaatgaaaa
      241 ctgtcgacaa gctctgtccg tttgctggca actgcaaggg tctcaatcct atttgtaatt
      301 attgaataat aaaacaatta taaatgtcaa atttgttttt tattaacgat acaaaccaaa
      361 cgcaacaaga acatttgtag tattatctat aattgaaaac gcgtagttat aatcgctgag
      421 gtaatattta aaatcatttt caaatgattc acagttaatt tgcgacaata taattttatt
      481 ttcacataaa ctagacgcct tgtcgtcttc ttcttcgtat tccttctctt tttcattttt
      541 ctcttcataa aaattaacat agttattatc gtatccatat atgtatctat cgtatagagt
      601 aaattttttg ttgtcataaa tatatatgtc ttttttaatg gggtgtatag taccgctgcg
      661 catagttttt ctgtaattta caacagtgct attttctggt agttcttcgg agtgtgttgc
      721 tttaattatt aaatttatat aatcaatgaa tttgggatcg tcggttttgt acaatatgtt
      781 gccggcatag tacgcagctt cttctagttc aattacacca ttttttagca gcaccggatt
      841 aacataactt tccaaaatgt tgtacgaacc gttaaacaaa aacagttcac ctcccttttc
      901 tatactattg tctgcgagca gttgtttgtt gttaaaaata acagccattg taatgagacg
      961 cacaaactaa tatcacaaac tggaaatgtc tatcaatata tagttgctga tggccggcct
     1021 attaatagta atcaattacg gggtcattag ttcatagccc atatatggag ttccgcgtta
     1081 cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc ccattgacgt
     1141 caataatgac gtatgttccc atagtaacgc caatagggac tttccattga cgtcaatggg
     1201 tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat atgccaagtc
     1261 cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc cagtacatga
     1321 ccttacggga ctttcctact tggcagtaca tctacgtatt agtcatcgct attaccatgc
     1381 tgatgcggtt ttggcagtac accaatgggc gtggatagcg gtttgactca cggggatttc
     1441 caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat caacgggact
     1501 ttccaaaatg tcgtaataac cccgccccgt tgacgcaaat gggcggtagg cgtgtacggt
     1561 gggaggtcta tataagcaga cgtcgtttag tgaaccgtca gatcactaga tgctttattg
     1621 cggtagttta tcacagttaa attgctaacg ccagtctcga acttaacgtg cagaagttgg
     1681 tcgtgaggca ctgggcaggt aagtatcggg ccctttgtgc ggggggagcg gctcggggct
     1741 gtccgcgggg ggacggctgc cttcgggggg gacggggcag ggcggggttc ggcttctggc
     1801 gtgtgaccgg cggctctaga gcctctgcta accatgttca tgccttcttc tttttcctac
     1861 agctcctggg caacgtgctg gttattgtgc tgtctcatca ttttggcaaa gaattggatc
     1921 ggaccgaaat taatacgact cactataggg gaattgtgag cggataacaa ttccccggag
     1981 ttaatccggg acctttaatt caacccaaca caatatatta tagttaaata agaattatta
     2041 tcaaatcatt tgtatattaa ttaaaatact atactgtaaa ttacatttta tttacaatca
     2101 aaggagatat accatggcac accatcacca ccatcacgga tctgtgagca agggcgagga
     2161 ggataacatg gccatcatca aggagttcat gcgcttcaag gtgcacatgg agggctccgt
     2221 gaacggccac gagttcgaga tcgagggcga gggcgagggc cgcccctacg agggcaccca
     2281 gaccgccaag ctgaaggtga ccaagggtgg ccccctgccc ttcgcctggg acatcctgtc
     2341 ccctcagttc atgtacggct ccaaggccta cgtgaagcac cccgccgaca tccccgacta
     2401 cttgaagctg tccttccccg agggcttcaa gtgggagcgc gtgatgaact tcgaggacgg
     2461 cggcgtggtg accgtgaccc aggactcctc cctgcaggac ggcgagttca tctacaaggt
     2521 gaagctgcgc ggcaccaact tcccctccga cggccccgta atgcagaaga agaccatggg
     2581 ctgggaggcc tcctccgagc ggatgtaccc cgaggacggc gccctgaagg gcgagatcaa
     2641 gcagaggctg aagctgaagg acggcggcca ctacgacgct gaggtcaaga ccacctacaa
     2701 ggccaagaag cccgtgcagc tgcccggcgc ctacaacgtc aacatcaagt tggacatcac
     2761 ctcccacaac gaggactaca ccatcgtgga acagtacgaa cgcgccgagg gccgccactc
     2821 caccggcggc atggacgagc tgtacaaggg atccttggct actacacttg aacgtattga
     2881 gaagaacttt gtcattactg acccaagatt gccagataat cccattatat tcgcgtccga
     2941 tagtttcttg cagttgacag aatatagccg tgaagaaatt ttgggaagaa actgcaggtt
     3001 tctacaaggt cctgaaactg atcgcgcgac agtgagaaaa attagagatg ccatagataa
     3061 ccaaacagag gtcactgttc agctgattaa ttatacaaag agtggtaaaa agttctggaa
     3121 cctctttcac ttgcagccta tgcgagatca gaagggagat gtccagtact ttattggggt
     3181 tcagttggat ggaactgagc atgtccgaga tgctgccgag agagagggag tcatgctgat
     3241 taagaaaact gcagaaaata ttgatgaggc ggcaaaagaa cttaagcttg aatcacacag
     3301 ctcatggtat gctcctgagg ccctgcagaa gtggctgcag ctgacccatg aggtggaggt
     3361 gcagtactac aacatcaaga agcaaaatgc agagaggcag ctgctggtgg ccaaggaggg
     3421 ggctgagaaa ataaaaaaga agagaaacac gctttttggt accttccatg tggcccacag
     3481 ctcttccctg gatgatgtgg atcataaaat cctaactgct aagcaagctc tgagtgaggt
     3541 gacagcggca ctgagggagc gcctgcaccg gtggcagcag atcgagatcc tctgcggttt
     3601 ccagattgtc aataaccccg gcatccactc cttggtggct gctctcaaca tcgaccccag
     3661 ctggatgggc agcacccgcc ctaaccccgc ccacttcatc atgactgacg atgtggatga
     3721 catggatgag gagattgtgt cgtaactcga gcaccaccat caccatcacc atcactaagt
     3781 gattaacctc aggtgcaggc tgcctatcag aaggtggtgg ctggtgtggc caatgccctg
     3841 gctcacaaat accactgaga tcgatctttt tccctctgcc aaaaattatg gggacatcat
     3901 gaagcccctt gagcatctga cttctggcta ataaaggaaa tttattttca ttgcaatagt
     3961 gtgttggaat tttttgtgtc tctcactcgg aaggacatat gggagggcaa atcatttaaa
     4021 acatcagaat gagtatttgg tttagagttt ggcaacatat gcccatatgt aactagcata
     4081 accccttggg gcctctaaac gggtcttgag gggttttttg ctgaaagcat gcggaggaaa
     4141 ttctccttga agtttccctg gtgttcaaag taaaggagtt tgcaccagac gcacctctgt
     4201 tcactggtcc ggcgtattaa aacacgatac attgttatta gtacatttat taagcgctag
     4261 attctgtgcg ttgttgattt acagacaatt gttgtacgta ttttaataat tcattaaatt
     4321 tataatcttt agggtggtat gttagagcga aaatcaaatg attttcagcg tctttatatc
     4381 tgaatttaaa tattaaatcc tcaatagatt tgtaaaatag gtttcgatta gtttcaaaca
     4441 agggttgttt ttccgaaccg atggctggac tatctaatgg attttcgctc aacgccacaa
     4501 aacttgccaa atcttgtagc agcaatctag ctttgtcgat attcgtttgt gttttgtttt
     4561 gtaataaagg ttcgacgtcg ttcaaaatat tatgcgcttt tgtatttctt tcatcactgt
     4621 cgttagtgta caattgactc gacgtaaaca cgttaaatag agcttggaca tatttaacat
     4681 cgggcgtgtt agctttatta ggccgattat cgtcgtcgtc ccaaccctcg tcgttagaag
     4741 ttgcttccga agacgatttt gccatagcca cacgacgcct attaattgtg tcggctaaca
     4801 cgtccgcgat caaatttgta gttgagcttt ttggaattgc gatcgcataa cttcgtatag
     4861 catacattat acgaagttat aagctcggaa cgctgcgctc ggtcgttcgg ctgcggcgag
     4921 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     4981 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     5041 tggcgttttt ccataggctc cgccccctga cgagcatcac aaaaatcgac gctcaagtca
     5101 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     5161 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     5221 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     5281 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     5341 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     5401 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     5461 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
     5521 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     5581 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     5641 tcctttgtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     5701 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     5761 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     5821 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     5881 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     5941 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     6001 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     6061 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     6121 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     6181 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     6241 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     6301 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     6361 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     6421 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     6481 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     6541 atcagggtta ttgtctcatg tccgcgcgtt
//
