LOCUS       Exported                6091 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Mammalian expression of the super-dark state of LOV2.
ACCESSION   .
VERSION     .
KEYWORDS    pTriEx-mCherry-LOVSD
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6091)
  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 6091)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6091
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     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..1509
                     /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             3234..3266
                     /codon_start=1
                     /product="HSV (herpes simplex virus) epitope tag"
                     /label=HSV tag
                     /translation="QPELAPEDPED"
     CDS             3273..3296
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     primer_bind     complement(3386..3405)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3451..3506
                     /label=-beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3505..3524)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     terminator      3594..3641
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     misc_recomb     3654..4359
                     /label=baculovirus recombination region (ORF1629)
                     /note="contains part of ORF1629"
     3'UTR           3742..4114
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     CDS             complement(3771..4359)
                     /codon_start=2
                     /product="baculovirus capsid-associated protein"
                     /label=ORF1629
                     /note="required for viral replication"
                     /translation="IPKSSTTNLIADVLADTINRRRVAMAKSSSEATSNDEGWDDDDNR
                     PNKANTPDVKYVQALFNVFTSSQLYTNDSDERNTKAHNILNDVEPLLQNKTQTNIDKAR
                     LLLQDLASFVALSENPLDSPAIGSEKQPLFETNRNLFYKSIEDLIFKFRYKDAENHLIF
                     ALTYHPKDYKFNELLKYVQQLSVNQQRTESSA"
     protein_bind    4368..4401
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     rep_origin      complement(4569..5156)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4648..4667)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5169..6029)
                     /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     5792..5811
                     /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 cccccggggc tactacactt gaacgtattg agaagaactt tgtcattact gacccaagat
     2821 tgccagataa tcccattata ttcgcgtccg atagtttctt gcagttgaca gaatatagcc
     2881 gtgaagaaat tttgggaaga aacgccaggt ttctacaagg tcctgaaact gatcgcgcga
     2941 cagtgagaaa aattagagat gccatagata accaaacaga ggtcactgtt cagctgatta
     3001 attatacaaa gagtggtaaa aagttctgga acctctttca cttgcagcct atgcgagatc
     3061 agaagggaga tgtccagtac tttattgggg ttcagaagga tggaactgag catgtccgag
     3121 atgctgccga gagagaggca gtcatggaga ttaagaaaac tgcagaagaa attgatgagg
     3181 cggcaaaaga actttaaaag cttgcggccg cacagctgta tacacgtgca agccagccag
     3241 aactcgcccc ggaagacccc gaggatctcg agcaccacca tcaccatcac catcactaag
     3301 tgattaacct caggtgcagg ctgcctatca gaaggtggtg gctggtgtgg ccaatgccct
     3361 ggctcacaaa taccactgag atcgatcttt ttccctctgc caaaaattat ggggacatca
     3421 tgaagcccct tgagcatctg acttctggct aataaaggaa atttattttc attgcaatag
     3481 tgtgttggaa ttttttgtgt ctctcactcg gaaggacata tgggagggca aatcatttaa
     3541 aacatcagaa tgagtatttg gtttagagtt tggcaacata tgcccatatg taactagcat
     3601 aaccccttgg ggcctctaaa cgggtcttga ggggtttttt gctgaaagca tgcggaggaa
     3661 attctccttg aagtttccct ggtgttcaaa gtaaaggagt ttgcaccaga cgcacctctg
     3721 ttcactggtc cggcgtatta aaacacgata cattgttatt agtacattta ttaagcgcta
     3781 gattctgtgc gttgttgatt tacagacaat tgttgtacgt attttaataa ttcattaaat
     3841 ttataatctt tagggtggta tgttagagcg aaaatcaaat gattttcagc gtctttatat
     3901 ctgaatttaa atattaaatc ctcaatagat ttgtaaaata ggtttcgatt agtttcaaac
     3961 aagggttgtt tttccgaacc gatggctgga ctatctaatg gattttcgct caacgccaca
     4021 aaacttgcca aatcttgtag cagcaatcta gctttgtcga tattcgtttg tgttttgttt
     4081 tgtaataaag gttcgacgtc gttcaaaata ttatgcgctt ttgtatttct ttcatcactg
     4141 tcgttagtgt acaattgact cgacgtaaac acgttaaata gagcttggac atatttaaca
     4201 tcgggcgtgt tagctttatt aggccgatta tcgtcgtcgt cccaaccctc gtcgttagaa
     4261 gttgcttccg aagacgattt tgccatagcc acacgacgcc tattaattgt gtcggctaac
     4321 acgtccgcga tcaaatttgt agttgagctt tttggaattg cgatcgcata acttcgtata
     4381 gcatacatta tacgaagtta taagctcgga acgctgcgct cggtcgttcg gctgcggcga
     4441 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     4501 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     4561 ctggcgtttt tccataggct ccgccccctg acgagcatca caaaaatcga cgctcaagtc
     4621 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     4681 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     4741 cgggaagcgt ggcgctttct caatgctcac gctgtaggta tctcagttcg gtgtaggtcg
     4801 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     4861 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     4921 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     4981 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     5041 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     5101 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     5161 atcctttgtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     5221 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     5281 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     5341 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     5401 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     5461 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     5521 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     5581 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     5641 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     5701 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     5761 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     5821 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     5881 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     5941 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     6001 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     6061 tatcagggtt attgtctcat gtccgcgcgt t
//
