LOCUS       sequence-157005-        6142 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of the dark state of LOV2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6142)
  AUTHORS   Wang H, Vilela M, Winkler A, Tarnawski M, Schlichting I, Yumerefendi
            H, Kuhlman B, Liu R, Danuser G, Hahn KM
  TITLE     LOVTRAP: an optogenetic system for photoinduced protein 
            dissociation.
  JOURNAL   Nat Methods. 2016 Jul 18. doi: 10.1038/nmeth.3926.
  PUBMED    27427858
REFERENCE   2  (bases 1 to 6142)
  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: "Nat 
            Methods. 2016 Jul 18. doi: 10.1038/nmeth.3926."
COMMENT     Sequence Label: pTriEx-mCherry-LOV2 C450A
FEATURES             Location/Qualifiers
     source          1..6142
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_recomb     110..937
                     /label=baculovirus recombination region (lef2/ORF603)
                     /note="contains ORF603 and part of lef2"
     CDS             110..234
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(272..877)
                     /codon_start=1
                     /product="baculovirus ORF603 protein"
                     /label=ORF603
                     /translation="MAVIFNNKQLLADNSIEKGGELFLFNGSYNILESYVNPVLLKNGV
                     IELEEAAYYAGNILYKTDDPKFIDYINLIIKATHSEELPENSTVVNYRKTMRSGTIHPI
                     KKDIYIYDNKKFTLYDRYIYGYDNNYVNFYEEKNEKEKEYEEEDDKASSLCENKIILSQ
                     INCESFENDFKYYLSDYNYAFSIIDNTTNVLVAFGLYR"
     enhancer        1005..1308
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1310..1512
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1463..1483
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1715..1737
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1799..1818
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     primer_bind     1860..1879
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1860..1878
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1879..1903
                     /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        1919..2028
                     /label=p10 promoter
                     /note="baculovirus promoter for expression in insect cells"
     CDS             2043..2750
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(2188..2206)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     CDS             2817..3245
                     /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
                     NARFLQGPETDRATVRKIRDAIDNQTEVTVQLINYTKSGKKFWNLFHLQPMRDQKGDVQ
                     YFIGVQLDGTEHVRDAAEREGVMLIKKTAENIDEAAKEL"
     CDS             3285..3317
                     /codon_start=1
                     /product="HSV (herpes simplex virus) epitope tag"
                     /label=HSV tag
                     /translation="QPELAPEDPED"
     CDS             3324..3347
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     primer_bind     complement(3437..3456)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3502..3557
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3556..3575)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     terminator      3645..3692
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     misc_recomb     3705..4410
                     /label=baculovirus recombination region (ORF1629)
                     /note="contains part of ORF1629"
     3'UTR           3793..4165
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     CDS             complement(3822..4410)
                     /codon_start=2
                     /product="baculovirus capsid-associated protein"
                     /label=ORF1629
                     /note="required for viral replication"
                     /translation="IPKSSTTNLIADVLADTINRRRVAMAKSSSEATSNDEGWDDDDNR
                     PNKANTPDVKYVQALFNVFTSSQLYTNDSDERNTKAHNILNDVEPLLQNKTQTNIDKAR
                     LLLQDLASFVALSENPLDSPAIGSEKQPLFETNRNLFYKSIEDLIFKFRYKDAENHLIF
                     ALTYHPKDYKFNELLKYVQQLSVNQQRTESSA"
     protein_bind    complement(4419..4452)
                     /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(4620..5207)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4699..4718)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5220..6011,6012..6080))
                     /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     5843..5862
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 aactgttaca ttgcaaacgt ggtttcgtgt gccaagtgtg aaaaccgatg tttaatcaag
       61 gctctgacgc atttctacaa ccacgactcc aagtgtgtgg gtgaagtcat gcatctttta
      121 atcaaatccc aagatgtgta taaaccacca aactgccaaa aaatgaaaac tgtcgacaag
      181 ctctgtccgt ttgctggcaa ctgcaagggt ctcaatccta tttgtaatta ttgaataata
      241 aaacaattat aaatgtcaaa tttgtttttt attaacgata caaaccaaac gcaacaagaa
      301 catttgtagt attatctata attgaaaacg cgtagttata atcgctgagg taatatttaa
      361 aatcattttc aaatgattca cagttaattt gcgacaatat aattttattt tcacataaac
      421 tagacgcctt gtcgtcttct tcttcgtatt ccttctcttt ttcatttttc tcttcataaa
      481 aattaacata gttattatcg tatccatata tgtatctatc gtatagagta aattttttgt
      541 tgtcataaat atatatgtct tttttaatgg ggtgtatagt accgctgcgc atagtttttc
      601 tgtaatttac aacagtgcta ttttctggta gttcttcgga gtgtgttgct ttaattatta
      661 aatttatata atcaatgaat ttgggatcgt cggttttgta caatatgttg ccggcatagt
      721 acgcagcttc ttctagttca attacaccat tttttagcag caccggatta acataacttt
      781 ccaaaatgtt gtacgaaccg ttaaacaaaa acagttcacc tcccttttct atactattgt
      841 ctgcgagcag ttgtttgttg ttaaaaataa cagccattgt aatgagacgc acaaactaat
      901 atcacaaact ggaaatgtct atcaatatat agttgctgat ggccggccta ttaatagtaa
      961 tcaattacgg ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg
     1021 gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg
     1081 tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta
     1141 cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtcc gccccctatt
     1201 gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac cttacgggac
     1261 tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatgct gatgcggttt
     1321 tggcagtaca ccaatgggcg tggatagcgg tttgactcac ggggatttcc aagtctccac
     1381 cccattgacg tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt
     1441 cgtaataacc ccgccccgtt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat
     1501 ataagcagac gtcgtttagt gaaccgtcag atcactagat gctttattgc ggtagtttat
     1561 cacagttaaa ttgctaacgc cagtctcgaa cttaacgtgc agaagttggt cgtgaggcac
     1621 tgggcaggta agtatcgggc cctttgtgcg gggggagcgg ctcggggctg tccgcggggg
     1681 gacggctgcc ttcggggggg acggggcagg gcggggttcg gcttctggcg tgtgaccggc
     1741 ggctctagag cctctgctaa ccatgttcat gccttcttct ttttcctaca gctcctgggc
     1801 aacgtgctgg ttattgtgct gtctcatcat tttggcaaag aattggatcg gaccgaaatt
     1861 aatacgactc actatagggg aattgtgagc ggataacaat tccccggagt taatccggga
     1921 cctttaattc aacccaacac aatatattat agttaaataa gaattattat caaatcattt
     1981 gtatattaat taaaatacta tactgtaaat tacattttat ttacaatcaa aggagatata
     2041 ccatggtgag caagggcgag gaggataaca tggccatcat caaggagttc atgcgcttca
     2101 aggtgcacat ggagggctcc gtgaacggcc acgagttcga gatcgagggc gagggcgagg
     2161 gccgccccta cgagggcacc cagaccgcca agctgaaggt gaccaagggt ggccccctgc
     2221 ccttcgcctg ggacatcctg tcccctcagt tcatgtacgg ctccaaggcc tacgtgaagc
     2281 accccgccga catccccgac tacttgaagc tgtccttccc cgagggcttc aagtgggagc
     2341 gcgtgatgaa cttcgaggac ggcggcgtgg tgaccgtgac ccaggactcc tccctgcagg
     2401 acggcgagtt catctacaag gtgaagctgc gcggcaccaa cttcccctcc gacggccctg
     2461 taatgcagaa gaagactatg ggctgggagg cctcctccga gcggatgtac cccgaggacg
     2521 gcgccctgaa gggcgagatc aagcagaggc tgaagctgaa ggacggcggc cactacgacg
     2581 ctgaggtcaa gaccacctac aaggccaaga agcccgtgca gctgcccggc gcctacaacg
     2641 tcaacatcaa gttggacatc acctcccaca acgaggacta caccatcgtg gaacagtacg
     2701 aacgcgccga gggccgccac tccaccggcg gcatggacga gctgtacaag ggttctggat
     2761 ccggtggttc tggtggttct ggtggttctg gtggttctgg tggttctggt ggtaccttgg
     2821 ctactacact tgaacgtatt gagaagaact ttgtcattac tgacccaaga ttgccagata
     2881 atcccattat attcgcgtcc gatagtttct tgcagttgac agaatatagc cgtgaagaaa
     2941 ttttgggaag aaacgccagg tttctacaag gtcctgaaac tgatcgcgcg acagtgagaa
     3001 aaattagaga tgccatagat aaccaaacag aggtcactgt tcagctgatt aattatacaa
     3061 agagtggtaa aaagttctgg aacctctttc acttgcagcc tatgcgagat cagaagggag
     3121 atgtccagta ctttattggg gttcagttgg atggaactga gcatgtccga gatgctgccg
     3181 agagagaggg agtcatgctg attaagaaaa ctgcagaaaa tattgatgag gcggcaaaag
     3241 aactttaaaa gcttgcggcc gcacagctgt atacacgtgc aagccagcca gaactcgccc
     3301 cggaagaccc cgaggatctc gagcaccacc atcaccatca ccatcactaa gtgattaacc
     3361 tcaggtgcag gctgcctatc agaaggtggt ggctggtgtg gccaatgccc tggctcacaa
     3421 ataccactga gatcgatctt tttccctctg ccaaaaatta tggggacatc atgaagcccc
     3481 ttgagcatct gacttctggc taataaagga aatttatttt cattgcaata gtgtgttgga
     3541 attttttgtg tctctcactc ggaaggacat atgggagggc aaatcattta aaacatcaga
     3601 atgagtattt ggtttagagt ttggcaacat atgcccatat gtaactagca taaccccttg
     3661 gggcctctaa acgggtcttg aggggttttt tgctgaaagc atgcggagga aattctcctt
     3721 gaagtttccc tggtgttcaa agtaaaggag tttgcaccag acgcacctct gttcactggt
     3781 ccggcgtatt aaaacacgat acattgttat tagtacattt attaagcgct agattctgtg
     3841 cgttgttgat ttacagacaa ttgttgtacg tattttaata attcattaaa tttataatct
     3901 ttagggtggt atgttagagc gaaaatcaaa tgattttcag cgtctttata tctgaattta
     3961 aatattaaat cctcaataga tttgtaaaat aggtttcgat tagtttcaaa caagggttgt
     4021 ttttccgaac cgatggctgg actatctaat ggattttcgc tcaacgccac aaaacttgcc
     4081 aaatcttgta gcagcaatct agctttgtcg atattcgttt gtgttttgtt ttgtaataaa
     4141 ggttcgacgt cgttcaaaat attatgcgct tttgtatttc tttcatcact gtcgttagtg
     4201 tacaattgac tcgacgtaaa cacgttaaat agagcttgga catatttaac atcgggcgtg
     4261 ttagctttat taggccgatt atcgtcgtcg tcccaaccct cgtcgttaga agttgcttcc
     4321 gaagacgatt ttgccatagc cacacgacgc ctattaattg tgtcggctaa cacgtccgcg
     4381 atcaaatttg tagttgagct ttttggaatt gcgatcgcat aacttcgtat agcatacatt
     4441 atacgaagtt ataagctcgg aacgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     4501 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     4561 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     4621 ttccataggc tccgccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     4681 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     4741 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     4801 tggcgctttc tcaatgctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     4861 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     4921 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     4981 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     5041 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
     5101 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     5161 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttgt
     5221 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     5281 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     5341 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     5401 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     5461 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     5521 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     5581 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     5641 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     5701 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     5761 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     5821 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     5881 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     5941 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     6001 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     6061 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     6121 tattgtctca tgtccgcgcg tt
//
