LOCUS Exported 7939 bp ds-DNA circular SYN 17-FEB-2022 DEFINITION Flp-dependent expression of mNaChBac-T2A-tdTomato. ACCESSION . VERSION . KEYWORDS pAAV-EF1-alpha-F-FLEX-mNaChBac-T2A-tdTomato SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7939) AUTHORS Xue M, Atallah BV, Scanziani M TITLE Equalizing excitation-inhibition ratios across visual cortical neurons. JOURNAL Nature. 2014 Jul 31;511(7511):596-600. doi: 10.1038/nature13321. Epub 2014 Jun 22. PUBMED 25043046 REFERENCE 2 (bases 1 to 7939) AUTHORS . TITLE Direct Submission JOURNAL Exported Feb 17, 2022 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7939 /organism="synthetic DNA construct" /mol_type="other DNA" repeat_region 1..141 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" repeat_region 12..141 /label=AAV2 ITR promoter 242..1419 /label=EF-1-alpha promoter /note="strong constitutive promoter for human elongation factor EF-1-alpha" intron 473..1410 /label=EF-1-alpha intron A /note="intron upstream of the start codon of human EF-1-alpha" primer_bind 1367..1387 /label=EF1a-F /note="Human elongation factor-1a promoter, forward primer" protein_bind 1510..1543 /label=FRT (minimal) /bound_moiety="FLP recombinase from the Saccharomyces cerevisiae 2u plasmid" /note="supports FLP-mediated excision but not integration (Turan and Bode, 2011)" CDS complement(1550..2980) /codon_start=1 /product="tandem dimeric (pseudo-monomeric) derivative of DsRed (Shaner et al., 2004)" /label=tdTomato /note="mammalian codon-optimized" /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH HLFLGHGTGSTGSGSSGTASSEDNNMAVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPY EGTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVM NFEDGGLVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGV LKGEIHQALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYE RSEGRHHLFLYGMDELYK" regulatory 2977..2986 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS complement(2996..3049) /codon_start=1 /product="2A peptide from Thosea asigna virus capsid protein" /label=T2A /note="Eukaryotic ribosomes fail to insert a peptide bond between the Gly and Pro residues, yielding separate polypeptides." /translation="EGRGSLLTCGDVEENPGP" protein_bind complement(4006..4039) /label=FRT (minimal) /bound_moiety="FLP recombinase from the Saccharomyces cerevisiae 2u plasmid" /note="supports FLP-mediated excision but not integration (Turan and Bode, 2011)" misc_feature 4066..4654 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(4119..4139) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(4537..4548) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" polyA_signal 4686..5162 /label=hGH poly(A) signal /note="human growth hormone polyadenylation signal" primer_bind complement(4885..4904) /label=hGH-PA-R /note="Human growth hormone terminator, reverse primer" repeat_region 5202..5342 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" repeat_region 5202..5331 /label=AAV2 ITR rep_origin 5417..5872 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(5504..5523) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 5714..5735 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(5889..5908) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 6008..6030 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(6068..6086) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 6154..6258 /gene="bla" /label=AmpR promoter CDS 6259..7119 /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 complement(6477..6496) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 7290..7878 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 7779..7798 /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc 61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca 121 actccatcac taggggttcc tgcggccgca cgcgtaagct ttgcaaagat ggataaagtt 181 ttaaacagag aggaatcttt gcagctaatg gaccttctag gtcttgaaag gagtgggaat 241 tggctccggt gcccgtcagt gggcagagcg cacatcgccc acagtccccg agaagttggg 301 gggaggggtc ggcaattgaa ccggtgccta gagaaggtgg cgcggggtaa actgggaaag 361 tgatgtcgtg tactggctcc gcctttttcc cgagggtggg ggagaaccgt atataagtgc 421 agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac aggtaagtgc 481 cgtgtgtggt tcccgcgggc ctggcctctt tacgggttat ggcccttgcg tgccttgaat 541 tacttccact ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg 601 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg 661 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct 721 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg 781 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc 841 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga 901 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct 961 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca 1021 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg 1081 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg 1141 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat 1201 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg 1261 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa 1321 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca 1381 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gtaccgaagt tcctattcta 1441 tcagaagtat aggaacttcg cagaatggta gctggattgt agctgctatt agcaatatga 1501 aacctcttag aagttcctat tctctagaaa gtataggaac ttcgaattct tacttgtaca 1561 gctcgtccat gccgtacagg aacaggtggt ggcggccctc ggagcgctcg tactgttcca 1621 cgatggtgta gtcctcgttg tgggaggtga tgtccagctt ggtgtccacg tagtagtagc 1681 cgggcagttg cacgggcttc ttggccatgt agatggtctt gaactccacc aggtagtggc 1741 cgccgtcctt cagcttcagg gcctggtgga tctcgccctt cagcacgccg tcgcgggggt 1801 acaggcgctc ggtggaggcc tcccagccca tggtcttctt ctgcattacg gggccgtcgg 1861 gggggaagtt ggtgccgcgc atcttcacct tgtagatcag cgtgccgtcc tgcagggagg 1921 agtcctgggt cacggtcacc agaccgccgt cctcgaagtt catcacgcgc tcccacttga 1981 agccctcggg gaaggacagc ttcttgtaat cggggatgtc ggcggggtgc ttcacgtacg 2041 ccttggagcc gtacatgaac tggggggaca ggatgtccca ggcgaagggc agggggccgc 2101 ccttggtcac cttcagcttg gcggtctggg tgccctcgta ggggcggccc tcgccctcgc 2161 cctcgatctc gaactcgtgg ccgttcatgg agccctccat gcgcaccttg aagcgcatga 2221 actctttgat gacggccatg ttgttgtcct cggaggaggc ggtgccggag ctgccgctgc 2281 cggtgctgcc ggtgccatgc cccaggaaca ggtggtggcg gccctcggag cgctcgtact 2341 gttccacgat ggtgtagtcc tcgttgtggg aggtgatgtc cagcttggtg tccacgtagt 2401 agtagccggg cagttgcacg ggcttcttgg ccatgtagat ggtcttgaac tccaccaggt 2461 agtggccgcc gtccttcagc ttcagggcct ggtggatctc gcccttcagc acgccgtcgc 2521 gggggtacag gcgctcggtg gaggcctccc agcccatggt cttcttctgc attacggggc 2581 cgtcgggggg gaagttggtg ccgcgcatct tcaccttgta gatcagcgtg ccgtcctgca 2641 gggaggagtc ctgggtcacg gtcaccagac cgccgtcctc gaagttcatc acgcgctccc 2701 acttgaagcc ctcggggaag gacagcttct tgtaatcggg gatgtcggcg gggtgcttca 2761 cgtacgcctt ggagccgtac atgaactggg gggacaggat gtcccaggcg aagggcaggg 2821 ggccgccctt ggtcaccttc agcttggcgg tctgggtgcc ctcgtagggg cggccctcgc 2881 cctcgccctc gatctcgaac tcgtggccgt tcatggagcc ctccatgcgc accttgaagc 2941 gcatgaactc tttgatgacc tcctcgccct tgctcaccat ggtggcggga tccgctgggc 3001 caggattctc ctcgacgtca ccgcatgtta gcagacttcc tctgccctct ccactgccga 3061 tatccttaag ttgtttgagc aagctcttca gttcggctac atcctttcgc aatgctgaaa 3121 tctcctgttt aagtccgtcg gcctccccat cttcctcatt gtccgtaagc tccgcctttt 3181 ccacgttgtt cacaatgact ccgatgaaaa gattgaaaat gataaaagta ccaatcagga 3241 caaacgagac aaagtagagc catgaccagg gcacctctgc aaaaatgggc cgcattacac 3301 cgctggccca gctctccaag gtcaccactt ggaacagcgt cagcaacgaa agctggaggt 3361 tcccaaagta ttcaggcgat acgtgctgga agagcattgt cccgatcact gcaaagatat 3421 agaagaagat ggacatcaag atcaagatat tgcccaacgc cggaatcgtc atcaccagcg 3481 catctacgag tctgcgcagg gaggggacga cggagatcgc cctgagtacc cgcagcactc 3541 taaggattcg caatactgtc acgaactggg ctcccgcgaa gatatgccca gccgcgacaa 3601 tcagaaaatc aaaccaattc cacgagctgc ggaagaaggc cgactttggg ttcgacgcca 3661 aaaacctcat cgcgatctcg atggtgaaga tccaaagcag gacgaggtca atccgataaa 3721 agagccactt gtggtccgcg tagatcctcg gatatgtttc gattcccacg atcagtgcat 3781 taaagaggat cagggcaatg acggtgaaag taaacgctct atggttgaca attttctgca 3841 tctttgacgt gaatgagttc tgtttctggc gtgcttccat tttcatggtg gcggctagct 3901 acgtaggcgc gccttaatta agaagttcct atacttctga tagaatagga acttcttgcc 3961 ttaacccaga aattatcact gttattcttt agaatggtgc aaagagaagt tcctatactt 4021 tctagagaat aggaacttcc agaattcgat atcaagctta tcgataatca acctctggat 4081 tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt 4141 ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc 4201 tcctccttgt ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg 4261 caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc 4321 accacctgtc agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa 4381 ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat 4441 tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc tgctcgccta tgttgccacc 4501 tggattctgc gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt 4561 ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag 4621 acgagtcgga tctccctttg ggccgcctcc ccgcatcgat accgagcgct gctcgagaga 4681 tctacgggtg gcatccctgt gacccctccc cagtgcctct cctggccctg gaagttgcca 4741 ctccagtgcc caccagcctt gtcctaataa aattaagttg catcattttg tctgactagg 4801 tgtccttcta taatattatg gggtggaggg gggtggtatg gagcaagggg caagttggga 4861 agacaacctg tagggcctgc ggggtctatt gggaaccaag ctggagtgca gtggcacaat 4921 cttggctcac tgcaatctcc gcctcctggg ttcaagcgat tctcctgcct cagcctcccg 4981 agttgttggg attccaggca tgcatgacca ggctcagcta atttttgttt ttttggtaga 5041 gacggggttt caccatattg gccaggctgg tctccaactc ctaatctcag gtgatctacc 5101 caccttggcc tcccaaattg ctgggattac aggcgtgaac cactgctccc ttccctgtcc 5161 ttctgatttt gtaggtaacc acgtgcggac cgagcggccg caggaacccc tagtgatgga 5221 gttggccact ccctctctgc gcgctcgctc gctcactgag gccgggcgac caaaggtcgc 5281 ccgacgcccg ggctttgccc gggcggcctc agtgagcgag cgagcgcgca gctgcctgca 5341 ggggcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatacg 5401 tcaaagcaac catagtacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt 5461 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc 5521 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct 5581 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga tttgggtgat 5641 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc 5701 acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc tatctcgggc 5761 tattcttttg atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg 5821 atttaacaaa aatttaacgc gaattttaac aaaatattaa cgtttacaat tttatggtgc 5881 actctcagta caatctgctc tgatgccgca tagttaagcc agccccgaca cccgccaaca 5941 cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg 6001 accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga 6061 cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct 6121 tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc 6181 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa 6241 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt 6301 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct 6361 gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc 6421 cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta 6481 tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac 6541 tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc 6601 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac 6661 ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg 6721 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac 6781 gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc 6841 gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt 6901 gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga 6961 gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc 7021 cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag 7081 atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca 7141 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc 7201 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca 7261 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc 7321 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta 7381 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt 7441 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc 7501 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg 7561 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg 7621 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag 7681 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc 7741 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat 7801 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg 7861 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc 7921 tggccttttg ctcacatgt //