LOCUS Exported 7291 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Expresses Cry2-mCherry-PKA W196R/F327A in mammalian cells.. ACCESSION . VERSION . KEYWORDS optoPKAW196R/F327A SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7291) AUTHORS O'Banion CP, Priestman MA, Hughes RM, Herring LE, Capuzzi SJ, Lawrence DS TITLE Design and Profiling of a Subcellular Targeted Optogenetic cAMP-Dependent Protein Kinase. JOURNAL Cell Chem Biol. 2017 Oct 25. pii: S2451-9456(17)30355-0. doi: 10.1016/j.chembiol.2017.09.011. PUBMED 29104065 REFERENCE 2 (bases 1 to 7291) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7291 /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" CDS 625..2118 /codon_start=1 /product="N-terminal photolyase homology region of Arabidopsis thaliana cryptochrome 2" /label=CRY2PHR /note="binds to CIB1 upon exposure to blue light (Kennedy et al., 2010)" /translation="MKMDKKTIVWFRRDLRIEDNPALAAAAHEGSVFPVFIWCPEEEGQ FYPGRASRWWMKQSLAHLSQSLKALGSDLTLIKTHNTISAILDCIRVTGATKVVFNHLY DPVSLVRDHTVKEKLVERGISVQSYNGDLLYEPWEIYCEKGKPFTSFNSYWKKCLDMSI ESVMLPPPWRLMPITAAAEAIWACSIEELGLENEAEKPSNALLTRAWSPGWSNADKLLN EFIEKQLIDYAKNSKKVVGNSTSLLSPYLHFGEISVRHVFQCARMKQIIWARDKNSEGE ESADLFLRGIGLREYSRYICFNFPFTHEQSLLSHLRFFPWDADVDKFKAWRQGRTGYPL VDAGMRELWATGWMHNRIRVIVSSFAVKFLLLPWKWGMKYFWDTLLDADLECDILGWQY ISGSIPDGHELDRLDNPALQGAKYDPEGEYIRQWLPELARLPTEWIHHPWDAPLTVLKA SGVELGTNYAKPIVDIDTARELLAKAISRTREAQIMIGAA" regulatory 2137..2146 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 2143..2850 /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(2288..2306) /label=mCherry-R /note="mCherry, reverse primer" primer_bind 2556..2575 /label=mCherry-F /note="mCherry, forward primer" polyA_signal 4079..4200 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(4116..4135) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 4170..4189 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin complement(4207..4662) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4344..4365) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 4556..4575 /label=F1ori-R /note="F1 origin, reverse primer" promoter 4689..4793 /gene="bla" /label=AmpR promoter promoter 4795..5152 /label=SV40 promoter /note="SV40 enhancer and early promoter" primer_bind complement(4818..4838) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" rep_origin 5003..5138 /label=SV40 ori /note="SV40 origin of replication" primer_bind 5065..5084 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 5187..5981 /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(5241..5260) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 5851..5870 /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind complement(6169..6188) /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" polyA_signal 6213..6260 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" rep_origin 6589..7177 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 7078..7097 /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 ccggactcag atctcgaggc caccatgaag atggacaaaa agactatagt ttggtttaga 661 agagacctaa ggattgagga taatcctgca ttagcagcag ctgctcacga aggatctgtt 721 tttcctgtct tcatttggtg tcctgaagaa gaaggacagt tttatcctgg aagagcttca 781 agatggtgga tgaaacaatc acttgctcac ttatctcaat ccttgaaggc tcttggatct 841 gacctcactt taatcaaaac ccacaacacg atttcagcga tcttggattg tatccgcgtt 901 accggtgcta caaaagtcgt ctttaaccac ctctatgatc ctgtttcgtt agttcgggac 961 cataccgtaa aggagaagct ggtggaacgt gggatctctg tgcaaagcta caatggagat 1021 ctattgtatg aaccgtggga gatatactgc gaaaagggca aaccttttac gagtttcaat 1081 tcttactgga agaaatgctt agatatgtcg attgaatccg ttatgcttcc tcctccttgg 1141 cggttgatgc caataactgc agcggctgaa gcgatttggg cgtgttcgat tgaagaacta 1201 gggctggaga atgaggccga gaaaccgagc aatgcgttgt taactagagc ttggtctcca 1261 ggatggagca atgctgataa gttactaaat gagttcatcg agaagcagtt gatagattat 1321 gcaaagaaca gcaagaaagt tgttgggaat tctacttcac tactttctcc gtatctccat 1381 ttcggggaaa taagcgtcag acacgttttc cagtgtgccc ggatgaaaca aattatatgg 1441 gcaagagata agaacagtga aggagaagaa agtgcagatc tttttcttag gggaatcggt 1501 ttaagagagt attctcggta tatatgtttc aacttcccgt ttactcacga gcaatcgttg 1561 ttgagtcatc ttcggttttt cccttgggat gctgatgttg ataagttcaa ggcctggaga 1621 caaggcagga ccggttatcc gttggtggat gccggaatga gagagctttg ggctaccgga 1681 tggatgcata acagaataag agtgattgtt tcaagctttg ctgtgaagtt tcttctcctt 1741 ccatggaaat ggggaatgaa gtatttctgg gatacacttt tggatgctga tttggaatgt 1801 gacatccttg gctggcagta tatctctggg agtatccccg atggccacga gcttgatcgc 1861 ttggacaatc ccgcgttaca aggcgccaaa tatgacccag aaggtgagta cataaggcaa 1921 tggcttcccg agcttgcgag attgccaact gaatggatcc atcatccatg ggacgctcct 1981 ttaaccgtac tcaaagcttc tggtgtggaa ctcggaacaa actatgcgaa acccattgta 2041 gacatcgaca cagctcgtga gctactagct aaagctattt caagaacccg tgaagcacag 2101 atcatgatcg gagcagcagc ccgggatcca ccggtcgcca ccatggtgag caagggcgag 2161 gaggataaca tggccatcat caaggagttc atgcgcttca aggtgcacat ggagggctcc 2221 gtgaacggcc acgagttcga gatcgagggc gagggcgagg gccgccccta cgagggcacc 2281 cagaccgcca agctgaaggt gaccaagggt ggccccctgc ccttcgcctg ggacatcctg 2341 tcccctcagt tcatgtacgg ctccaaggcc tacgtgaagc accccgccga catccccgac 2401 tacttgaagc tgtccttccc cgagggcttc aagtgggagc gcgtgatgaa cttcgaggac 2461 ggcggcgtgg tgaccgtgac ccaggactcc tccctgcagg acggcgagtt catctacaag 2521 gtgaagctgc gcggcaccaa cttcccctcc gacggccccg taatgcagaa gaagaccatg 2581 ggctgggagg cctcctccga gcggatgtac cccgaggacg gcgccctgaa gggcgagatc 2641 aagcagaggc tgaagctgaa ggacggcggc cactacgacg ctgaggtcaa gaccacctac 2701 aaggccaaga agcccgtgca gctgcccggc gcctacaacg tcaacatcaa gttggacatc 2761 acctcccaca acgaggacta caccatcgtg gaacagtacg aacgcgccga gggccgccac 2821 tccaccggcg gcatggacga gctgtacaag agtgctggtg gtagtgctgg tggtagtgct 2881 ggtggtagtg ctggtggtat gggcaacgcc gccgccgcca agaagggcag cgagcaggag 2941 agcgtgaaag agttcctagc caaagccaag gaagatttcc tgaaaaaatg ggagacccct 3001 tctcagaata cagcccagtt ggatcagttt gatagaatca agacccttgg caccggctcc 3061 tttgggcgag tgatgctggt gaagcacaag gagagtggga accactacgc catgaagatc 3121 ttagacaagc agaaggtggt gaagctaaag cagatcgagc acactctgaa tgagaagcgc 3181 atcctgcagg ccgtcaactt cccgttcctg gtcaaacttg aattctcctt caaggacaac 3241 tcaaacctgt atatggtcat ggagtatgta gctggtggcg agatgttctc ccacctacgg 3301 cggattggaa ggttcagcga gccccatgcc cgtttctacg cggcgcagat cgtcctgacc 3361 tttgagtatc tgcactccct ggacctcatc taccgggacc tgaagcccga gaatcttctc 3421 atcgaccagc agggctatat tcaggtgaca gacttcggtt ttgccaagcg tgtgaaaggc 3481 cgtactcgga ccttgtgtgg gacccctgag tacttggccc ccgagattat cctgagcaaa 3541 ggctacaaca aggctgtgga ctggtgggct ctcggagtcc tcatctacga gatggctgct 3601 ggttacccac ccttcttcgc tgaccagcct atccagatct atgagaaaat cgtctctggg 3661 aaggtgcggt tcccatccca cttcagctct gacttgaagg acctgctgcg gaaccttctg 3721 caagtggatc tcaccaagcg ctttgggaac ctcaagaacg gggtcaatga catcaagaac 3781 cacaagtggt ttgccacgac tgactggatt gccatctatc agagaaaggt ggaagctccc 3841 ttcataccaa agtttaaagg ccctggggac acgagtaacg ccgacgacta tgaggaggaa 3901 gagatccggg tctccatcaa tgagaagtgt ggcaaggagt ttactgagtt ttaatgagcg 3961 gccgcgactc tagatcataa tcagccatac cacatttgta gaggttttac ttgctttaaa 4021 aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg ttgttgttaa 4081 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa 4141 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta 4201 aggcgtaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca 4261 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga 4321 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg 4381 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat 4441 caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag 4501 ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga 4561 agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa 4621 ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcaggtggc acttttcggg 4681 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc 4741 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtcctgagg 4801 cggaaagaac cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc 4861 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc 4921 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat 4981 agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc 5041 gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga 5101 gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca aagatcgatc 5161 aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc 5221 cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct 5281 ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg 5341 acctgtccgg tgccctgaat gaactgcaag acgaggcagc gcggctatcg tggctggcca 5401 cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc 5461 tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga 5521 aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc 5581 cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc 5641 ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg 5701 ccaggctcaa ggcgagcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct 5761 gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc 5821 tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc 5881 ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc 5941 agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga 6001 aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt 6061 ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg 6121 cggggatctc atgctggagt tcttcgccca ccctaggggg aggctaactg aaacacggaa 6181 ggagacaata ccggaaggaa cccgcgctat gacggcaata aaaagacaga ataaaacgca 6241 cggtgttggg tcgtttgttc ataaacgcgg ggttcggtcc cagggctggc actctgtcga 6301 taccccaccg agaccccatt ggggccaata cgcccgcgtt tcttcctttt ccccacccca 6361 ccccccaagt tcgggtgaag gcccagggct cgcagccaac gtcggggcgg caggccctgc 6421 catagcctca ggttactcat atatacttta gattgattta aaacttcatt tttaatttaa 6481 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt 6541 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt 6601 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg 6661 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca 6721 gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt 6781 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga 6841 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc 6901 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact 6961 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga 7021 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg 7081 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt 7141 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt 7201 acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga 7261 ttctgtggat aaccgtatta ccgccatgca t //