LOCUS Exported 7144 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION expresses a Cre-dependent red light-shifted channelrhodopsin variant. ACCESSION . VERSION . KEYWORDS pAAV-EF1-alpha-DIO-ReaChR-P2A-dTomato SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7144) AUTHORS Messier JE, Chen H, Cai ZL, Xue M TITLE Targeting light-gated chloride channels to neuronal somatodendritic domain reduces their excitatory effect in the axon. JOURNAL Elife. 2018 Aug 9;7. pii: 38506. doi: 10.7554/eLife.38506. PUBMED 30091701 REFERENCE 2 (bases 1 to 7144) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7144 /organism="synthetic DNA construct" /mol_type="other DNA" protein_bind 35..68 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." protein_bind 119..152 /label=lox2272 /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GGATACTT). lox2272 sites are compatible with each other, but incompatible with loxP or loxN sites (Lee and Saito, 1988)." regulatory 162..171 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 1233..1289 /codon_start=1 /product="2A peptide from porcine teschovirus-1 polyprotein" /label=P2A /note="Eukaryotic ribosomes fail to insert a peptide bond between the Gly and Pro residues, yielding separate polypeptides." /translation="ATNFSLLKQAGDVEENPGP" CDS 1290..1991 /codon_start=1 /product="dimeric variant of DsRed fluorescent protein (Shaner et al., 2004)" /label=dTomato /note="mammalian codon-optimized" /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH HLFLYGMDELYK" primer_bind complement(1423..1441) /label=mCherry-R /note="mCherry, reverse primer" primer_bind 1691..1710 /label=mCherry-F /note="mCherry, forward primer" primer_bind 1885..1908 /label=DsRed1-C /note="DsRed1, forward primer" protein_bind complement(2009..2042) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." protein_bind complement(2093..2126) /label=lox2272 /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GGATACTT). lox2272 sites are compatible with each other, but incompatible with loxP or loxN sites (Lee and Saito, 1988)." promoter complement(2157..3335) /label=EF-1-alpha promoter /note="strong constitutive promoter for human elongation factor EF-1-alpha" intron 2166..3104 /label=EF-1-alpha intron A /note="intron upstream of the start codon of human EF-1-alpha" primer_bind complement(2189..2209) /label=EF1a-F /note="Human elongation factor-1a promoter, forward primer" repeat_region 3436..3565 /label=AAV2 ITR (alternate) /note="Functional equivalent of wild-type AAV2 ITR" rep_origin complement(3627..4215) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(3707..3726) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(4386..5246) /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 5009..5028 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(5247..5351) /gene="bla" /label=AmpR promoter primer_bind 5419..5437 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" primer_bind complement(5475..5497) /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind 5597..5616 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" rep_origin complement(5633..6088) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(5770..5791) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 5982..6001 /label=F1ori-R /note="F1 origin, reverse primer" repeat_region 6163..6303 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" repeat_region 6174..6303 /label=AAV2 ITR polyA_signal 6421..6532 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" misc_feature 6566..10 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" CDS 6672..6683 /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" primer_bind 7081..7101 /label=WPRE-R /note="WPRE, reverse primer" ORIGIN 1 gaggttgatt atcgataagc ttgatatcga attcataact tcgtatagca tacattatac 61 gaagttattc tttgcaccat tctaaagaat aacagtgata atttctgggt taaggcaaat 121 aacttcgtat aggatacttt atacgaagtt atgctagcgg cgccaccatg gtgagcagaa 181 gaccctggct gctggccctg gccctggccg tggccctggc cgccggcagc gccggcgcca 241 gcaccggcag cgacgccacc gtgcccgtgg ccacccagga cggccccgac tacgtgttcc 301 acagagccca cgagagaatg ctgttccaga ccagctacac cctggagaac aacggcagcg 361 tgatctgcat ccccaacaac ggccagtgct tctgcctggc ctggctgaag agtaacggca 421 ccaacgccga gaagctggcc gccaacatcc tgcagtgggt ggtgtttgcg ctgagcgtgg 481 cgtgcctggg ctggtatgcg tatcaggcgt ggcgcgcgac ctgcggctgg gaagaagtgt 541 atgtggcgct gattgaaatg atgaaaagca ttattgaagc gtttcatgaa tttgatagcc 601 cggcgaccct gtggctgagc agcggcaacg gcgtggtgtg gatgcgctat ggcgaatggc 661 tgctgacctg ccccgtgatt ctgattcatc tgagcaacct gaccggcctg aaagatgatt 721 atagcaaacg caccatgggc ctgctggtga gcgacgtggg ctgcattgtg tggggcgcga 781 ccagcgcgat gtgcaccggc tggaccaaaa ttctgttttt tctgattagc ctgagctatg 841 gcatgtatac ctattttcat gcggccaaag tgtatattga agcgtttcat accgtgccga 901 aaggcctgtg cagacagctg gtgagagcca tggcctggct gttcttcgtg agctggggca 961 tgttccccgt gctgttcctg ctgggccccg agggcttcgg ccatattagc ccgtatggca 1021 gcgcgattgg ccatagcatt ctggatctga ttgcgaagaa catgtggggc gtgctgggca 1081 actatctgcg cgtgaaaatt catgaacata ttctgctgta tggcgatatt cgcaaaaaac 1141 agaaaattac cattgcgggc caggaaatgg aagtggaaac cctggtggcg gaagaagaag 1201 ataagtacga gagcagcgtt aacggaagcg gagccactaa cttctccctg ttgaaacaag 1261 caggggatgt cgaagagaat cccgggccaa tggtgagcaa gggcgaggag gtcatcaaag 1321 agttcatgcg cttcaaggtg cgcatggagg gctccatgaa cggccacgag ttcgagatcg 1381 agggcgaggg cgagggccgc ccctacgagg gcacccagac cgccaagctg aaggtgacca 1441 agggcggccc cctgcccttc gcctgggaca tcctgtcccc ccagttcatg tacggctcca 1501 aggcgtacgt gaagcacccc gccgacatcc ccgattacaa gaagctgtcc ttccccgagg 1561 gcttcaagtg ggagcgcgtg atgaacttcg aggacggcgg tctggtgacc gtgacccagg 1621 actcctccct gcaggacggc acgctgatct acaaggtgaa gatgcgcggc accaacttcc 1681 cccccgacgg ccccgtaatg cagaagaaga ccatgggctg ggaggcctcc accgagcgcc 1741 tgtacccccg cgacggcgtg ctgaagggcg agatccacca ggccctgaag ctgaaggacg 1801 gcggccacta cctggtggag ttcaagacca tctacatggc caagaagccc gtgcaactgc 1861 ccggctacta ctacgtggac accaagctgg acatcacctc ccacaacgag gactacacca 1921 tcgtggaaca gtacgagcgc tccgagggcc gccaccacct gttcctgtac ggcatggacg 1981 agctgtacaa gtaagtcgac ggcgcgccat aacttcgtat aatgtatgct atacgaagtt 2041 attaagaggt ttcatattgc taatagcagc tacaatccag ctaccattct gcataacttc 2101 gtataaagta tcctatacga agttattccg gagtcgactc tagaggatcc ggtacctcac 2161 gacacctgaa atggaagaaa aaaactttga accactgtct gaggcttgag aatgaaccaa 2221 gatccaaact caaaaagggc aaattccaag gagaattaca tcaagtgcca agctggccta 2281 acttcagtct ccacccactc agtgtgggga aactccatcg cataaaaccc ctccccccaa 2341 cctaaagacg acgtactcca aaagctcgag aactaatcga ggtgcctgga cggcgcccgg 2401 tactccgtgg agtcacatga agcgacggct gaggacggaa aggccctttt cctttgtgtg 2461 ggtgactcac ccgcccgctc tcccgagcgc cgcgtcctcc attttgagct ccctgcagca 2521 gggccgggaa gcggccatct ttccgctcac gcaactggtg ccgaccgggc cagccttgcc 2581 gcccagggcg gggcgataca cggcggcgcg aggccaggca ccagagcagg ccggccagct 2641 tgagactacc cccgtccgat tctcggtggc cgcgctcgca ggccccgcct cgccgaacat 2701 gtgcgctggg acgcacgggc cccgtcgccg cccgcggccc caaaaaccga aataccagtg 2761 tgcagatctt ggcccgcatt tacaagacta tcttgccaga aaaaaagcgt cgcagcaggt 2821 catcaaaaat tttaaatggc tagagactta tcgaaagcag cgagacaggc gcgaaggtgc 2881 caccagattc gcacgcggcg gccccagcgc ccaggccagg cctcaactca agcacgaggc 2941 gaaggggctc cttaagcgca aggcctcgaa ctctcccacc cacttccaac ccgaagctcg 3001 ggatcaagaa tcacgtactg cagccaggtg gaagtaattc aaggcacgca agggccataa 3061 cccgtaaaga ggccaggccc gcgggaacca cacacggcac ttacctgtgt tctggcggca 3121 aacccgttgc gaaaaagaac gttcacggcg actactgcac ttatatacgg ttctccccca 3181 ccctcgggaa aaaggcggag ccagtacacg acatcacttt cccagtttac cccgcgccac 3241 cttctctagg caccggttca attgccgacc cctcccccca acttctcggg gactgtgggc 3301 gatgtgcgct ctgcccactg acgggcaccg gagccaattc ccactccttt caagacctag 3361 aaggtccatt agctgcaaag attcctctct gtttaaaact ttatccatct ttgcaaagct 3421 tacgcgtgcg gccgcaggaa cccctagtga tggagttggc cactccctct ctgcgcgctc 3481 gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg cccgggcggc ctcagtgagc 3541 gagcgagcgc gcagctgcct gcaggacatg tgagcaaaag gccagcaaaa ggccaggaac 3601 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac 3661 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg 3721 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac 3781 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat 3841 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag 3901 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac 3961 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt 4021 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt 4081 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc 4141 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga 4201 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac 4261 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc 4321 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct 4381 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca 4441 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct 4501 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca 4561 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc 4621 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg 4681 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct 4741 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa 4801 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta 4861 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc 4921 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg 4981 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa 5041 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg 5101 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc 5161 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg 5221 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat 5281 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata 5341 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac cattattatc 5401 atgacattaa cctataaaaa taggcgtatc acgaggccct ttcgtctcgc gcgtttcggt 5461 gatgacggtg aaaacctctg acacatgcag ctcccggaga cggtcacagc ttgtctgtaa 5521 gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg 5581 ggctggctta actatgcggc atcagagcag attgtactga gagtgcacca taaaattgta 5641 aacgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac 5701 caatagaccg aaatcggcaa aatcccttat aaatcaaaag aatagcccga gatagagttg 5761 agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa 5821 gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc caaatcaagt 5881 tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt 5941 agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga 6001 gcgggcgcta aggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc 6061 gcgcttaatg cgccgctaca gggcgcgtac tatggttgct ttgacgtatg cggtgtgaaa 6121 taccgcacag atgcgtaagg agaaaatacc gcatcaggcg cccctgcagg cagctgcgcg 6181 ctcgctcgct cactgaggcc gcccgggcaa agcccgggcg tcgggcgacc tttggtcgcc 6241 cggcctcagt gagcgagcga gcgcgcagag agggagtggc caactccatc actaggggtt 6301 cctgcggccg ctcggtccgc acgtgagatc tcatgcctgc tattgtcttc ccaatcctcc 6361 cccttgctgt cctgccccac cccacccccc agaatagaat gacacctact cagacaatgc 6421 gatgcaattt cctcatttta ttaggaaagg acagtgggag tggcaccttc cagggtcaag 6481 gaaggcacgg gggaggggca aacaacagat ggctggcaac tagaaggcac agtcgaagat 6541 ctctcgagca gcgctcggta tcgatgcggg gaggcggccc aaagggagat ccgactcgtc 6601 tgagggcgaa ggcgaagacg cggaagaggc cgcagagccg gcagcaggcc gcgggaagga 6661 aggtccgctg gattgagggc cgaagggacg tagcagaagg acgtcccgcg cagaatccag 6721 gtggcaacat aggcgagcag ccaaggaaag gacgatgatt tccccgacaa caccacggaa 6781 ttgtcagtgc ccaacagccg agcccctgtc cagcagcggg caaggcaggc ggcgatgagt 6841 tccgccgtgg caatagggag ggggaaagcg aaagtcccgg aaaggagctg acaggtggtg 6901 gcaatgcccc aaccagtggg ggttgcgtca gcaaacacag tgcacaccac gccacgttgc 6961 ctgacaacgg gccacaactc ctcataaaga gacagcaacc aggatttata caaggaggag 7021 aaaatgaaag ccatacggga agcaatagca tgatacaaag gcattaaagc agcgtatcca 7081 catagcgtaa aaggagcaac atagttaaga ataccagtca atctttcaca aattttgtaa 7141 tcca //