| 质粒名称: | pAAV-CaMKIIa-hChR2(E123A)-mCherry |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 35506 |
| 存储实验室: | Karl Deisseroth |
|
载体骨架基本信息 |
|
| 载体名称: | pAAV |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | 5385 bp |
| 载体修饰: | 添加CaMKIIa促进剂和WPRE |
| 载体类型: | 哺乳细胞表达,AAV |
| 高拷贝/低拷贝: | 高拷贝 |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | hChR2 |
| 物种来源: | 莱茵衣藻 |
| 插入大小: | 1646 bp |
| 基因突变: | E123A |
| 启动子 | CaMKIIa |
| 标签/融合蛋白: | mCherry（插入件上的C端子） |
| 克隆方法: | 限制性内切酶 |
| 5' 克隆位点: | BamHI(未破坏) |
| 3' 克隆位点: | EcoRI (未破坏) |
| 5' 测序引物 | AGTCCTGCAGTATTGTGTAT |
| 3' 测序引物 | GCAATAGCATGATACAAAGG |
|
文献引用方法 |
|
| 材料和方法部分: | pAAV-CaMKIIa-hChR2(E123A)-mCherry was a gift from Karl Deisseroth (Addgene plasmid # 35506 ; http://n2t.net/addgene:35506 ; RRID:Addgene_35506) |
| 参考文献部分: | Principles for applying optogenetic tools derived from direct comparative analysis of microbial opsins. Mattis J, Tye KM, Ferenczi EA, Ramakrishnan C, O'Shea DJ, Prakash R, Gunaydin LA, Hyun M, Fenno LE, Gradinaru V, Yizhar O, Deisseroth K. Nat Methods. 2011 Dec 18;9(2):159-72. doi: 10.1038/nmeth.1808. 10.1038/nmeth.1808 PubMed 22179551 |
买家导航
载体基本信息
载体图谱质粒图谱和多克隆位点信息
-mCherry质粒图谱.png)
载体简介
载体序列
LOCUS pAAV-CaMKIIa-hChR2(E123A)-mCherry 7020 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION synthetic circular DNA
ACCESSION .
VERSION .
KEYWORDS pAAV-CaMKIIa-hChR2(E123A)-mCherry
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7020)
AUTHORS Mattis J, Tye KM, Ferenczi EA, Ramakrishnan C, O'Shea DJ, Prakash R,
Gunaydin LA, Hyun M, Fenno LE, Gradinaru V, Yizhar O, Deisseroth K
TITLE Principles for applying optogenetic tools derived from direct
comparative analysis of microbial opsins.
JOURNAL Nat Methods. 2011 Dec 18;9(2):159-72. doi: 10.1038/nmeth.1808.
PUBMED 22179551
REFERENCE 2 (bases 1 to 7020)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..7020
/organism="synthetic DNA construct"
/mol_type="other DNA"
repeat_region 54..183
/label=AAV2 ITR (alternate)
/note="Functional equivalent of wild-type AAV2 ITR"
regulatory 1523..1532
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 2468..3175
/codon_start=1
/product="monomeric derivative of DsRed fluorescent protein
(Shaner et al., 2004)"
/label=mCherry
/note="mammalian codon-optimized"
/translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
GRHSTGGMDELYK"
primer_bind complement(2610..2628)
/label=mCherry-R
/note="mCherry, reverse primer"
primer_bind 2878..2897
/label=mCherry-F
/note="mCherry, forward primer"
misc_feature 3200..3788
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
primer_bind complement(3253..3273)
/label=WPRE-R
/note="WPRE, reverse primer"
CDS complement(3671..3682)
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
polyA_signal 3820..4296
/label=hGH poly(A) signal
/note="human growth hormone polyadenylation signal"
primer_bind complement(4019..4038)
/label=hGH-PA-R
/note="Human growth hormone terminator, reverse primer"
repeat_region 4336..4476
/label=AAV2 ITR
/note="inverted terminal repeat of adeno-associated virus
serotype 2"
repeat_region 4336..4465
/label=AAV2 ITR
rep_origin 4551..5006
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(4638..4657)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 4848..4869
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(5023..5042)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 5142..5164
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind complement(5202..5220)
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
promoter 5288..5392
/gene="bla"
/label=AmpR promoter
CDS 5393..6253
/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(5611..5630)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 6424..7012
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 6913..6932
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
ORIGIN
1 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgtcctgcag
61 gcagctgcgc gctcgctcgc tcactgaggc cgcccgggcg tcgggcgacc tttggtcgcc
121 cggcctcagt gagcgagcga gcgcgcagag agggagtggc caactccatc actaggggtt
181 cctgcggccg cacgcgttta acattatggc cttaggtcac ttcatctcca tggggttctt
241 cttctgattt tctagaaaat gagatggggg tgcagagagc ttcctcagtg acctgcccag
301 ggtcacatca gaaatgtcag agctagaact tgaactcaga ttactaatct taaattccat
361 gccttggggg catgcaagta cgatatacag aaggagtgaa ctcattaggg cagatgacca
421 atgagtttag gaaagaagag tccagggcag ggtacatcta caccacccgc ccagccctgg
481 gtgagtccag ccacgttcac ctcattatag ttgcctctct ccagtcctac cttgacggga
541 agcacaagca gaaactggga caggagcccc aggagaccaa atcttcatgg tccctctggg
601 aggatgggtg gggagagctg tggcagaggc ctcaggaggg gccctgctgc tcagtggtga
661 cagatagggg tgagaaagca gacagagtca ttccgtcagc attctgggtc tgtttggtac
721 ttcttctcac gctaaggtgg cggtgtgata tgcacaatgg ctaaaaagca gggagagctg
781 gaaagaaaca aggacagaga cagaggccaa gtcaaccaga ccaattccca gaggaagcaa
841 agaaaccatt acagagacta caagggggaa gggaaggaga gatgaattag cttcccctgt
901 aaaccttaga acccagctgt tgccagggca acggggcaat acctgtctct tcagaggaga
961 tgaagttgcc agggtaacta catcctgtct ttctcaagga ccatcccaga atgtggcacc
1021 cactagccgt taccatagca actgcctctt tgccccactt aatcccatcc cgtctgttaa
1081 aagggcccta tagttggagg tgggggaggt aggaagagcg atgatcactt gtggactaag
1141 tttgttcgca tccccttctc caaccccctc agtacatcac cctgggggaa cagggtccac
1201 ttgctcctgg gcccacacag tcctgcagta ttgtgtatat aaggccaggg caaagaggag
1261 caggttttaa agtgaaaggc aggcaggtgt tggggaggca gttaccgggg caacgggaac
1321 agggcgtttc ggaggtggtt gccatgggga cctggatgct gacgaaggct cgcgaggctg
1381 tgagcagcca cagtgccctg ctcagaagcc ccaagctcgt cagtcaagcc ggttctccgt
1441 ttgcactcag gagcacgggc aggcgagtgg cccctagttc tgggggcagc tctagagcgg
1501 taccggatcc ccgggtaccg gtgccaccat ggattatgga ggcgctttgt ctgccgtcgg
1561 acgcgaactt ttgttcgtta ctaatcctgt ggtggtgaac gggtccgtcc tggtccctga
1621 ggatcaatgt tactgtgccg gatggattga atctcgcggc acgaacggcg ctcagaccgc
1681 gtcaaatgtc ctgcagtggc ttgcagcagg attcagcatt ttgctgctga tgttctatgc
1741 ctaccaaacc tggaaatcta catgcggctg ggaggagatc tatgtgtgcg ccattgaaat
1801 ggttaaggtg attctcgagt tcttttttga gtttaagaat ccctctatgc tctaccttgc
1861 cacaggacac cgggtgcagt ggctgcgcta tgcagcctgg ctgctcactt gtcctgtcat
1921 ccttatccac ctgagcaacc tcaccggcct gagcaacgac tacagcagga gaaccatggg
1981 actccttgtc tcagacatcg ggactatcgt gtggggggct accagcgcca tggcaaccgg
2041 ctatgttaaa gtcatcttct tttgtcttgg attgtgctat ggcgcgaaca cattttttca
2101 cgccgccaaa gcatatatcg agggttatca tactgtgcca aagggtcggt gccgccaggt
2161 cgtgaccggc atggcatggc tgtttttcgt gagctggggt atgttcccaa ttctcttcat
2221 tttggggccc gaaggttttg gcgtcctgag cgtctatggc tccaccgtag gtcacacgat
2281 tattgatctg atgagtaaaa attgttgggg gttgttggga cactacctgc gcgtcctgat
2341 ccacgagcac atattgattc acggagatat ccgcaaaacc accaaactga acatcggcgg
2401 aacggagatc gaggtcgaga ctctcgtcga agacgaagcc gaggccggag ccgtgccagc
2461 ggccgccgtg agcaagggcg aggaggataa catggccatc atcaaggagt tcatgcgctt
2521 caaggtgcac atggagggct ccgtgaacgg ccacgagttc gagatcgagg gcgagggcga
2581 gggccgcccc tacgagggca cccagaccgc caagctgaag gtgaccaagg gtggccccct
2641 gcccttcgcc tgggacatcc tgtcccctca gttcatgtac ggctccaagg cctacgtgaa
2701 gcaccccgcc gacatccccg actacttgaa gctgtccttc cccgagggct tcaagtggga
2761 gcgcgtgatg aacttcgagg acggcggcgt ggtgaccgtg acccaggact cctccctgca
2821 ggacggcgag ttcatctaca aggtgaagct gcgcggcacc aacttcccct ccgacggccc
2881 cgtaatgcag aagaagacca tgggctggga ggcctcctcc gagcggatgt accccgagga
2941 cggcgccctg aagggcgaga tcaagcagag gctgaagctg aaggacggcg gccactacga
3001 cgctgaggtc aagaccacct acaaggccaa gaagcccgtg cagctgcccg gcgcctacaa
3061 cgtcaacatc aagttggaca tcacctccca caacgaggac tacaccatcg tggaacagta
3121 cgaacgcgcc gagggccgcc actccaccgg cggcatggac gagctgtaca agtaagaatt
3181 cgatatcaag cttatcgata atcaacctct ggattacaaa atttgtgaaa gattgactgg
3241 tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta
3301 tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat cctggttgct
3361 gtctctttat gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt
3421 tgctgacgca acccccactg gttggggcat tgccaccacc tgtcagctcc tttccggaac
3481 tttcgctttc cccctcccta ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg
3541 ctggacaggg gctcggctgt tgggcactga caattccgtg gtgttgtcgg ggaaatcatc
3601 gtcctttcct tggctgctcg cctatgttgc cacctggatt ctgcgcggga cgtccttctg
3661 ctacgtccct tcggccctca atccagcgga ccttccttcc cgcggcctgc tgccggctct
3721 gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt cggatctccc tttgggccgc
3781 ctccccgcat cgataccgag cgctgctcga gagatctacg ggtggcatcc ctgtgacccc
3841 tccccagtgc ctctcctggc cctggaagtt gccactccag tgcccaccag ccttgtccta
3901 ataaaattaa gttgcatcat tttgtctgac taggtgtcct tctataatat tatggggtgg
3961 aggggggtgg tatggagcaa ggggcaagtt gggaagacaa cctgtagggc ctgcggggtc
4021 tattgggaac caagctggag tgcagtggca caatcttggc tcactgcaat ctccgcctcc
4081 tgggttcaag cgattctcct gcctcagcct cccgagttgt tgggattcca ggcatgcatg
4141 accaggctca gctaattttt gtttttttgg tagagacggg gtttcaccat attggccagg
4201 ctggtctcca actcctaatc tcaggtgatc tacccacctt ggcctcccaa attgctggga
4261 ttacaggcgt gaaccactgc tcccttccct gtccttctga ttttgtaggt aaccacgtgc
4321 ggaccgagcg gccgcaggaa cccctagtga tggagttggc cactccctct ctgcgcgctc
4381 gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg
4441 cctcagtgag cgagcgagcg cgcagctgcc tgcaggggcg cctgatgcgg tattttctcc
4501 ttacgcatct gtgcggtatt tcacaccgca tacgtcaaag caaccatagt acgcgccctg
4561 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
4621 cagcgcctta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
4681 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
4741 gcacctcgac cccaaaaaac ttgatttggg tgatggttca cgtagtgggc catcgccctg
4801 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
4861 ccaaactgga acaacactca actctatctc gggctattct tttgatttat aagggatttt
4921 gccgatttcg gtctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt
4981 taacaaaata ttaacgttta caattttatg gtgcactctc agtacaatct gctctgatgc
5041 cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg
5101 tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca
5161 gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt
5221 tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg
5281 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
5341 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat
5401 tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc
5461 tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg
5521 ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg
5581 ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga
5641 cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta
5701 ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc
5761 tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc
5821 gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg
5881 ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc
5941 aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca
6001 acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct
6061 tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat
6121 cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg
6181 gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat
6241 taagcattgg taactgtcag accaagttta ctcatatata ctttagattg atttaaaact
6301 tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat
6361 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
6421 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
6481 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
6541 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
6601 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
6661 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
6721 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
6781 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
6841 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
6901 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
6961 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
//
