pAAV-CAG-Fluc

载体基本信息

载体质粒图谱和多克隆位点信息

载体简介

This AAV is a fLuc construct driven by the ubiquitous CAG promoter.The roughly 1.8Kb CAG promoter (also known as CBA promoter or CAGGS promoter) is a strong synthetic promoter frequently used to drive high levels of gene expression in mammalian cells. The CAG promoter is composed of the following regulatory elements: (C) cytomegalovirus (CMV) early enhancer element; (A) the promoter region, the first exon, and the first intron of chicken beta-Actin gene, and (G) the splice acceptor of the rabbit beta-Globin gene. Like the EF1a promoter, the CAG promoter is commonly used as an alternative for the CMV promoter, from which the expression is decreased due to methylation/silencing. In many cell types tested, the CAG and EF1a promoters give much higher levels of expression than other commonly used cellular promoters such as the UBC and PGK promoters.Firefly luciferase (fLuc) is an oxidative enzyme used in bioluminescence and is distinct from a photoprotein. In the luciferase reaction, light is emitted when luciferase acts on the appropriate luciferin substrate. Photon emission can be detected by a light-sensitive apparatus such as a luminometer or modified optical microscope. Firefly luciferase is commonly used as a reporter to assess the transcriptional activity in cells.

载体序列

LOCUS KC152483 8323 bp DNA circular SYN 03-JUN-2013DEFINITION Expression vector pAAV-CAG-Fluc, complete sequence.ACCESSION KC152483REFERENCE 1 (bases 1 to 8323) AUTHORS Chen,Y., Cao,L., Luo,C., Ditzel,D.A., Peter,J. and Sprengel,R. TITLE RANGE: Gene Transfer of Reversibly Controlled Polycistronic Genes JOURNAL Mol Ther Nucleic Acids 2, E85 (2013) PUBMED 23571608 JOURNAL Submitted (08-NOV-2012) Molecular Neurobiology, Max Planck institute for medical research, Jahnstr. 29, Heidelberg, BW 69120, GermanyFEATURES Location/Qualifiers source 1..8323 /organism="Expression vector pAAV-CAG-Fluc" /mol_type="other DNA" /db_xref="taxon:1337866" repeat_region 19..162 /note="AAV2 ITR" /rpt_type=inverted /rpt_type=terminal regulatory 200..1140 /regulatory_class="promoter" /note="CMV earlyenhancer/chicken beta actin promoter: CAG" misc_feature 1157..2809 /note="firefly luciferase" misc_feature 2810..2863 /note="2A peptide from Thosea asigna virus" misc_feature 2876..3724 /note="tTSKid" misc_feature 2876..3508 /note="Tet repressor: tetR[B/E]" misc_feature 3509..3526 /note="nuclear localization signal: NLS" misc_feature 3533..3718 /note="KRAB (Kruppel-associated box) domain; from Rat zinc finger protein (kid-1)" misc_feature 3725..3778 /note="2A peptide from Thosea asigna virus" gene 3791..4447 /gene="mKO" CDS 3791..4447 /gene="mKO" /codon_start=1 /transl_table=11 /product="monomeric Kusabira-orange" /protein_id="AGM53878.1" /db_xref="GI:507593976" /translation="MVSVIKPEMKMRYYMDGSVNGHEFTIEGEGTGRPYEGHQEMTLR VTMAKGGPMPFAFDLVSHVFCYGHRPFTKYPEEIPDYFKQAFPEGLSWERSLEFEDGG SASVSAHISLRGNTFYHKSKFTGVNFPADGPIMQNQSVDWEPSTEKITASDGVLKGDV TMYLKLEGGGNHKCQFKTTYKAAKKILKMPGSHYISHRLVRKTEGNITELVEDAVAHS " ncRNA 4478..5074 /ncRNA_class="other" /note="post-transcriptional regulatory element: WPRE" misc_feature 5107..5317 /note="bovine growth hormone polyA signal: bGH pA" repeat_region complement(5327..5470) /note="AAV2 ITR" /rpt_type=inverted /rpt_type=terminal gene 6659..7321 /gene="Bla" CDS 6659..7321 /gene="Bla" /note="ampicillin resistance" /codon_start=1 /transl_table=11 /product="beta-lactamase" /protein_id="AGM53877.1" /db_xref="GI:507593975" /translation="MSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYSPVTEKH LTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRWEPELN EAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSALPAG WFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGASLIK HW"ORIGIN 1 gggggggggg ggggggggtt ggccactccc tctctgcgcg ctcgctcgct cactgaggcc 61 gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg cggcctcagt gagcgagcga 121 gcgcgcagag agggagtggc caactccatc actaggggtt cctagatcta ggatcacgcg 181 ttctagtcct agggatgtac cattgacgtc aataatgacg tatgttccca tagtaacgcc 241 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc 301 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg 361 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat 421 ctacgtatta gtcatcgcta ttaccatggt cgaggtgagc cccacgttct gcttcactct 481 ccccatctcc cccccctccc cacccccaat tttgtattta tttatttttt aattattttg 541 tgcagcgatg ggggcggggg gggggggggg gcgcgcgcca ggcggggcgg ggcggggcga 601 ggggcggggc ggggcgaggc ggagaggtgc ggcggcagcc aatcagagcg gcgcgctccg 661 aaagtttcct tttatggcga ggcggcggcg gcggcggccc tataaaaagc gaagcgcgcg 721 gcgggcggga gtcgctgcgc gctgccttcg ccccgtgccc cgctccgccg ccgcctcgcg 781 ccgcccgccc cggctctgac tgaccgcgtt actcccacag gtgagcgggc gggacggccc 841 ttctcctccg ggctgtaatt agcgcttggt ttaatgacgg cttgtttctt ttctgtggct 901 gcgtgaaagc cttgaggggc tccgggaggg ccctttgtgc ggggggagcg gctcggggct 961 gtccgcgggg ggacggctgc cttcgggggg gacggggcag ggcggggttc ggcttctggc 1021 gtgtgaccgg cggctctaga gcctctgcta accatgttca tgccttcttc tttttcctac 1081 agctcctggg caacgtgctg gttattgtgc tgtctcatca ttttggcaaa gaattggatc 1141 tagtcgattc gaattcatgg aagacgccaa aaacataaag aaaggcccgg cgccattcta 1201 tccgctagag gatggaaccg ctggagagca actgcataag gctatgaaga gatacgccct 1261 ggttcctgga acaattgctt ttacagatgc acatatcgag gtgaacatca cgtacgcgga 1321 atacttcgaa atgtccgttc ggttggcaga agctatgaaa cgatatgggc tgaatacaaa 1381 tcacagaatc gtcgtatgca gtgaaaactc tcttcaattc tttatgccgg tgttgggcgc 1441 gttatttatc ggagttgcag ttgcgcccgc gaacgacatt tataatgaac gtgaattgct 1501 caacagtatg aacatttcgc agcctaccgt agtgtttgtt tccaaaaagg ggttgcaaaa 1561 aattttgaac gtgcaaaaaa aattaccaat aatccagaaa attattatca tggattctaa 1621 aacggattac cagggatttc agtcgatgta cacgttcgtc acatctcatc tacctcccgg 1681 ttttaatgaa tacgattttg taccagagtc ctttgatcgt gacaaaacaa ttgcactgat 1741 aatgaactcc tctggatcta ctgggttacc taagggtgtg gcccttccgc atagaactgc 1801 ctgcgtcaga ttctcgcatg ccagagatcc tatttttggc aatcaaatca ttccggatac 1861 tgcgatttta agtgttgttc cattccatca cggttttgga atgtttacta cactcggata 1921 tttgatatgt ggatttcgag tcgtcttaat gtatagattt gaagaagagc tgtttttacg 1981 atcccttcag gattacaaaa ttcaaagtgc gttgctagta ccaaccctat tttcattctt 2041 cgccaaaagc actctgattg acaaatacga tttatctaat ttacacgaaa ttgcttctgg 2101 gggcgcacct ctttcgaaag aagtcgggga agcggttgca aaacgcttcc atcttccagg 2161 gatacgacaa ggatatgggc tcactgagac tacatcagct attctgatta cacccgaggg 2221 ggatgataaa ccgggcgcgg tcggtaaagt tgttccattt tttgaagcga aggttgtgga 2281 tctggatacc gggaaaacgc tgggcgttaa tcagagaggc gaattatgtg tcagaggacc 2341 tatgattatg tccggttatg taaacaatcc ggaagcgacc aacgccttga ttgacaagga 2401 tggatggcta cattctggag acatagctta ctgggacgaa gacgaacact tcttcatagt 2461 tgaccgcttg aagtctttaa ttaaatacaa aggataccag gtggcccccg ctgaattgga 2521 gtcgatattg ttacaacacc ccaacatctt cgacgcgggc gtggcaggtc ttcccgacga 2581 tgacgccggt gaacttcccg ccgccgttgt tgttttggag cacggaaaga cgatgacgga 2641 aaaagagatc gtggattacg tcgccagtca agtaacaacc gcgaaaaagt tgcgcggagg 2701 agttgtgttt gtggacgaag taccgaaagg tcttaccgga aaactcgacg caagaaaaat 2761 cagagagatc ctcataaagg ccaagaaggg cggaaagtcc aaattgctcg agggcagagg 2821 aagtcttcta acatgcggtg acgtggagga gaatcccggc cctgcaccgg gatccatgtc 2881 cagattagat aaaagtaaag tgattaacag cgcattagag ctgcttaatg aggtcggaat 2941 cgaaggttta acaacccgta aactcgccca gaagctaggt gtagagcagc ctacattgta 3001 ttggcacgtg cgcaacaagc agactcttat gaacatgctt tcagaggcaa tactggcgaa 3061 gcatcacacc cgttcagcac cgttaccgac tgagagttgg cagcagtttc tccaggaaaa 3121 tgctctgagt ttccgtaaag cattactggt ccatcgtgat ggagcccgat tgcatatagg 3181 gacctctcct acgccccccc agtttgaaca agcagaggcg caactacgct gtctatgcga 3241 tgcagggttt tcggtcgagg aggctctttt cattctgcaa tctatcagcc attttacgtt 3301 gggtgcagta ttagaggagc aagcaacaaa ccagatagaa aataatcatg tgatagacgc 3361 tgcaccacca ttattacaag aggcatttaa tattcaggcg agaacctctg ctgaaatggc 3421 cttccatttc gggctgaaat cattaatatt tggattttct gcacagttag atgaaaaaaa 3481 gcatacaccc attgaggatg gtaataaacc aaaaaagaag agaaagctag cagtgtcagt 3541 gacatttgaa gatgtggctg tgctctttac tcgggacgag tggaagaagc tggatctgtc 3601 tcagagaagc ctgtaccgtg aggtgatgct ggagaattac agcaacctgg cctccatggc 3661 aggattcctg tttaccaaac caaaggtgat ctccctgttg cagcaaggag aggacccgta 3721 tctcgagggc agaggaagtc ttctaacatg cggtgacgtg gaggagaatc ccggccctgc 3781 accgggatcc atggtgagtg tgattaaacc agaaatgaag atgaggtact acatggacgg 3841 ctccgtcaat gggcatgagt tcacaattga aggtgaaggc acaggcagac cttacgaggg 3901 acatcaagag atgacactac gcgtcacaat ggccaagggc gggccaatgc ctttcgcgtt 3961 tgacttagtg tcacacgtgt tctgttacgg ccacagacct tttactaaat atccagaaga 4021 gataccagac tatttcaaac aagcatttcc tgaaggcctg tcatgggaaa ggtcgttgga 4081 gttcgaagat ggtgggtccg cttcagtcag tgcgcatata agccttagag gaaacacctt 4141 ctaccacaaa tccaaattta ctggggttaa ctttcctgcc gatggtccta tcatgcaaaa 4201 ccaaagtgtt gattgggagc catcaaccga gaaaattact gccagcgacg gagttctgaa 4261 gggtgatgtt acgatgtacc taaaacttga aggaggcggc aatcacaaat gccaattcaa 4321 gactacttac aaggcggcaa aaaagattct taaaatgcca ggaagccatt acatcagcca 4381 tcgcctcgtc aggaaaaccg aaggcaacat tactgagctg gtagaagatg cagtagctca 4441 ttcctgatgt acaagtaaag cggccgcaag cttatcgata atcaacctct ggattacaaa 4501 atttgtgaaa gattgactgg tattcttaac tatgttgctc cttttacgct atgtggatac 4561 gctgctttaa tgcctttgta tcatgctatt gcttcccgta tggctttcat tttctcctcc 4621 ttgtataaat cctggttgct gtctctttat gaggagttgt ggcccgttgt caggcaacgt 4681 ggcgtggtgt gcactgtgtt tgctgacgca acccccactg gttggggcat tgccaccacc 4741 tgtcagctcc tttccgggac tttcgctttc cccctcccta ttgccacggc ggaactcatc 4801 gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt tgggcactga caattccgtg 4861 gtgttgtcgg ggaagctgac gtcctttcca tggctgctcg cctgtgttgc cacctggatt 4921 ctgcgcggga cgtccttctg ctacgtccct tcggccctca atccagcgga ccttccttcc 4981 cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt 5041 cggatctccc tttgggccgc ctccccgcct gatcgatacc gtcgactaga gctcgctgat 5101 cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt 5161 ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat 5221 cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg 5281 gggaggattg ggaagacaat agcaggcatg ctggggagag atctgaggaa cccctagtga 5341 tggagttggc cactccctct ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc 5401 ccgggcgtcg ggcgaccttt ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg 5461 gagtggccaa cccccccccc cccccccctg cagcctggcg taatagcgaa gaggcccgca 5521 ccgatcgccc ttcccaacag ttgcgtagcc tgaatggcga atggcgcgac gcgccctgta 5581 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca 5641 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct 5701 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc 5761 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat 5821 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc 5881 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc 5941 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta 6001 acaaaatatt aacgtttaca atttcctgat gcggtatttt ctccttacgc atctgtgcgg 6061 tatttcacac cgcatatggt gcactctcag tacaatctgc tctgatgccg catagttaag 6121 ccagccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc 6181 atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc 6241 gtcatcaccg aaacgcgcga gacgaaaggg cctcgtgata cgcctatttt tataggttaa 6301 tgtcatgata ataatggttt cttagacgtc aggtggcact tttcggggaa atgtgcgcgg 6361 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata 6421 accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg 6481 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac 6541 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact 6601 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat 6661 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga 6721 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac 6781 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat 6841 gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac 6901 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct 6961 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac 7021 gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac aattaataga 7081 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg 7141 gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact 7201 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac 7261 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta 7321 actgtcagac caagtttact catatatact ttagattgat ttaaaacttc atttttaatt 7381 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga 7441 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc 7501 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt 7561 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc 7621 gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc 7681 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg 7741 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg 7801 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga 7861 actgagatac ctacagcgtg agcattgaga aagcgccacg cttcccgaag ggagaaaggc 7921 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg 7981 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg 8041 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt 8101 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc 8161 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg 8221 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc 8281 gcctctcccc gcgcgttggc cgattcatta atgcaggctg cag