LOCUS Exported 7437 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Expresses mCherry tag, then IRES, then Arabidopsis CRY2 (with L348F mutation) fused to N-terminal fragment of Cre DNA recombinase. ACCESSION . VERSION . KEYWORDS CRY2(L348F)-CreN SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7437) AUTHORS Taslimi A, Zoltowski B, Miranda JG, Pathak GP, Hughes RM, Tucker CL TITLE Optimized second-generation CRY2-CIB dimerizers and photoactivatable Cre recombinase. JOURNAL Nat Chem Biol. 2016 Apr 11. doi: 10.1038/nchembio.2063. PUBMED 27065233 REFERENCE 2 (bases 1 to 7437) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7437 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 61..364 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 365..568 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 519..539 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" regulatory 607..616 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 613..1320 /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(758..776) /label=mCherry-R /note="mCherry, reverse primer" primer_bind 1026..1045 /label=mCherry-F /note="mCherry, forward primer" primer_bind complement(1922..1938) /label=KS primer /note="common sequencing primer, one of multiple similar variants" primer_bind complement(1923..1939) /label=pBluescriptKS /note="For pBluescript vector" CDS 1950..3785 /codon_start=1 /product="Arabidopsis thaliana cryptochrome 2" /label=CRY2 /note="binds to CIB1 upon exposure to blue light (Kennedy et al., 2010)" /translation="MKMDKKTIVWFRRDLRIEDNPALAAAAHEGSVFPVFIWCPEEEGQ FYPGRASRWWMKQSLAHLSQSLKALGSDLTLIKTHNTISAILDCIRVTGATKVVFNHLY DPVSLVRDHTVKEKLVERGISVQSYNGDLLYEPWEIYCEKGKPFTSFNSYWKKCLDMSI ESVMLPPPWRLMPITAAAEAIWACSIEELGLENEAEKPSNALLTRAWSPGWSNADKLLN EFIEKQLIDYAKNSKKVVGNSTSLLSPYLHFGEISVRHVFQCARMKQIIWARDKNSEGE ESADLFLRGIGLREYSRYICFNFPFTHEQSLLSHLRFFPWDADVDKFKAWRQGRTGYPL VDAGMREFWATGWMHNRIRVIVSSFAVKFLLLPWKWGMKYFWDTLLDADLECDILGWQY ISGSIPDGHELDRLDNPALQGAKYDPEGEYIRQWLPELARLPTEWIHHPWDAPLTVLKA SGVELGTNYAKPIVDIDTARELLAKAISRTREAQIMIGAAPDEIVADSFEALGANTIKE PGLCPSVSSNDQQVPSAVRYNGSKRVKPEEEEERDMKKSRGFDERELFSTAESSSSSSV FFVSQSCSLASEGKNLEGIQDSSDQITTSLGKNGCK" polyA_signal 4225..4346 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(4262..4281) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 4316..4335 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin complement(4353..4808) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4490..4511) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 4702..4721 /label=F1ori-R /note="F1 origin, reverse primer" promoter 4835..4939 /gene="bla" /label=AmpR promoter promoter 4941..5298 /label=SV40 promoter /note="SV40 enhancer and early promoter" primer_bind complement(4964..4984) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" rep_origin 5149..5284 /label=SV40 ori /note="SV40 origin of replication" primer_bind 5211..5230 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 5333..6127 /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 LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(5387..5406) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 5997..6016 /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind complement(6315..6334) /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" polyA_signal 6359..6406 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" rep_origin 6735..7323 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 7224..7243 /label=pBR322ori-F /note="pBR322 origin, forward primer" 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 gctagcgcta 601 ccggtcgcca ccatggtgag caagggcgag gaggataaca tggccatcat caaggagttc 661 atgcgcttca aggtgcacat ggagggctcc gtgaacggcc acgagttcga gatcgagggc 721 gagggcgagg gccgccccta cgagggcacc cagaccgcca agctgaaggt gaccaagggt 781 ggccccctgc ccttcgcctg ggacatcctg tcccctcagt tcatgtacgg ctccaaggcc 841 tacgtgaagc accccgccga catccccgac tacttgaagc tgtccttccc cgagggcttc 901 aagtgggagc gcgtgatgaa cttcgaggac ggcggcgtgg tgaccgtgac ccaggactcc 961 tccctgcagg acggcgagtt catctacaag gtgaagctgc gcggcaccaa cttcccctcc 1021 gacggccccg taatgcagaa gaagaccatg ggctgggagg cctcctccga gcggatgtac 1081 cccgaggacg gcgccctgaa gggcgagatc aagcagaggc tgaagctgaa ggacggcggc 1141 cactacgacg ctgaggtcaa gaccacctac aaggccaaga agcccgtgca gctgcccggc 1201 gcctacaacg tcaacatcaa gttggacatc acctcccaca acgaggacta caccatcgtg 1261 gaacagtacg aacgcgccga gggccgccac tccaccggcg gcatggacga gctgtacaag 1321 tgtggcggct agtactccgg tattgcggta cccttgtacg cctgttttat actcccttcc 1381 cgtaacttag acgcacaaaa ccaagttcaa tagaaggggg tacaaaccag taccaccacg 1441 aacaagcact tctgtttccc cggtgatgtc gtatagactg cttgcgtggt tgaaagcgac 1501 ggatccgtta tccgcttatg tacttcgaga agcccagtac cacctcggaa tcttcgatgc 1561 gttgcgctca gcactcaacc ccagagtgta gcttaggctg atgagtctgg acatccctca 1621 ccggtgacgg tggtccaggc tgcgttggcg gcctacctat ggctaacgcc atgggacgct 1681 agttgtgaac aaggtgtgaa gagcctattg agctacntaa gaatcctccg gcccctgaat 1741 gcggctaatc ccaacctcgg agcaggtggt cacaaaccag tgattggcct gtcgtaacgc 1801 gcaagtccgt ggcggaaccg actactttgg gtgtccgtgt ttccttttat tttattgtgg 1861 ctgcttatgg tgacaatcac agattgttat cataaagcga attggatagg atcaagctta 1921 tcgataccgt cgacctcgag ctcgccacca tgaagatgga caaaaagact atagtttggt 1981 ttagaagaga cctaaggatt gaggataatc ctgcattagc agcagctgct cacgaaggat 2041 ctgtttttcc tgtcttcatt tggtgtcctg aagaagaagg acagttttat cctggaagag 2101 cttcaagatg gtggatgaaa caatcacttg ctcacttatc tcaatccttg aaggctcttg 2161 gatctgacct cactttaatc aaaacccaca acacgatttc agcgatcttg gattgtatcc 2221 gcgttaccgg tgctacaaaa gtcgtcttta accacctcta tgatcctgtt tcgttagttc 2281 gggaccatac cgtaaaggag aagctggtgg aacgtgggat ctctgtgcaa agctacaatg 2341 gagatctatt gtatgaaccg tgggagatat actgcgaaaa gggcaaacct tttacgagtt 2401 tcaattctta ctggaagaaa tgcttagata tgtcgattga atccgttatg cttcctcctc 2461 cttggcggtt gatgccaata actgcagcgg ctgaagcgat ttgggcgtgt tcgattgaag 2521 aactagggct ggagaatgag gccgagaaac cgagcaatgc gttgttaact agagcttggt 2581 ctccaggatg gagcaatgct gataagttac taaatgagtt catcgagaag cagttgatag 2641 attatgcaaa gaacagcaag aaagttgttg ggaattctac ttcactactt tctccgtatc 2701 tccatttcgg ggaaataagc gtcagacacg ttttccagtg tgcccggatg aaacaaatta 2761 tatgggcaag agataagaac agtgaaggag aagaaagtgc agatcttttt cttaggggaa 2821 tcggtttaag agagtattct cggtatatat gtttcaactt cccgtttact cacgagcaat 2881 cgttgttgag tcatcttcgg tttttccctt gggatgctga tgttgataag ttcaaggcct 2941 ggagacaagg caggaccggt tatccgttgg tggatgccgg aatgagagag ttttgggcta 3001 ccggatggat gcataacaga ataagagtga ttgtttcaag ctttgctgtg aagtttcttc 3061 tccttccatg gaaatgggga atgaagtatt tctgggatac acttttggat gctgatttgg 3121 aatgtgacat ccttggctgg cagtatatct ctgggagtat ccccgatggc cacgagcttg 3181 atcgcttgga caatcccgcg ttacaaggcg ccaaatatga cccagaaggt gagtacataa 3241 ggcaatggct tcccgagctt gcgagattgc caactgaatg gatccatcat ccatgggacg 3301 ctcctttaac cgtactcaaa gcttctggtg tggaactcgg aacaaactat gcgaaaccca 3361 ttgtagacat cgacacagct cgtgagctac tagctaaagc tatttcaaga acccgtgaag 3421 cacagatcat gatcggagca gcacctgatg agattgtagc agatagcttc gaggccttag 3481 gggctaatac cattaaagaa cctggtcttt gcccatctgt gtcttctaat gaccaacaag 3541 taccttcggc tgttcgttac aacgggtcaa agagagtgaa acctgaggaa gaagaagaga 3601 gagacatgaa gaaatctagg ggattcgatg aaagggagtt gttttcgact gctgaatctt 3661 cttcttcttc gagtgtgttt ttcgtttcgc agtcttgctc gttggcatca gaagggaaga 3721 atctggaagg tattcaagat tcatctgatc agattactac aagtttggga aaaaatggtt 3781 gcaaaggtgg cggtggctct ggaggtggtg ggtccggagg aggcggccgc acgagtgatg 3841 aggttcgcaa gaacctgatg gacatgttca gggatcgcca ggcgttttct gagcatacct 3901 ggaaaatgct tctgtccgtt tgccggtcgt gggcggcatg gtgcaagttg aataaccgga 3961 aatggtttcc cgcagaacct gaagatgttc gcgattatct tctatatctt caggcgcgcg 4021 gtctggcagt aaaaactatc cagcaacatt tgggccagct aaacatgctt catcgtcggt 4081 ccgggctgta acccgggatc caccggatct agataactga tcataatcag ccataccaca 4141 tttgtagagg ttttacttgc tttaaaaaac ctcccacacc tccccctgaa cctgaaacat 4201 aaaatgaatg caattgttgt tgttaacttg tttattgcag cttataatgg ttacaaataa 4261 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt 4321 ttgtccaaac tcatcaatgt atcttaacgc gtaaattgta agcgttaata ttttgttaaa 4381 attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa 4441 aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa 4501 caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca 4561 gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg 4621 taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc 4681 ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc 4741 aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca 4801 gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt 4861 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata 4921 atattgaaaa aggaagagtc ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta 4981 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat 5041 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc 5101 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta 5161 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca 5221 gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga 5281 ggcctaggct tttgcaaaga tcgatcaaga gacaggatga ggatcgtttc gcatgattga 5341 acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga 5401 ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg 5461 gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaagacga 5521 ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt 5581 tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct 5641 gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct 5701 gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg 5761 agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca 5821 ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg agcatgcccg acggcgagga 5881 tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt 5941 ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt 6001 ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct 6061 ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt 6121 cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca 6181 cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg 6241 gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccaccct 6301 agggggaggc taactgaaac acggaaggag acaataccgg aaggaacccg cgctatgacg 6361 gcaataaaaa gacagaataa aacgcacggt gttgggtcgt ttgttcataa acgcggggtt 6421 cggtcccagg gctggcactc tgtcgatacc ccaccgagac cccattgggg ccaatacgcc 6481 cgcgtttctt ccttttcccc accccacccc ccaagttcgg gtgaaggccc agggctcgca 6541 gccaacgtcg gggcggcagg ccctgccata gcctcaggtt actcatatat actttagatt 6601 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc 6661 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag 6721 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa 6781 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg 6841 aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag 6901 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg 6961 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga 7021 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc 7081 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc 7141 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga 7201 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt 7261 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg 7321 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac 7381 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc catgcat //