| 质粒名称: | pAAV_hsyn_NES-his-CaMPARI2-WPRE-SV40 |
|---|---|
| 出品公司： | addgene |
| 目录编号: | 101060 |
| 存储实验室: | Eric Schreiter |
|
载体骨架基本信息 |
|
| 载体名称: | pAAV |
| 空载体大小: | 4267 bp |
| 完整质粒大小: | 5517bp |
| 载体类型: | AAV表达 |
| 载体抗性: | 氨苄霉素 |
| 生长温度: | 37 ℃ |
| 宿主菌: | NEB Stable |
| 拷贝数: | 高拷贝 |
|
插入基因信息 |
|
| 插入基因名称: | CaMPARI2/CaMPARI_Q142V-F198Y-C202T-L217I-N345S-W391F-V398L |
| 物种来源: | Synthetic |
| 插入大小: | 1446bp |
| 启动子: | hsyn (synapsin-1) |
| 标签/融合蛋白: |
NES_his (N terminal on insert)
FLAG-HA-myc (C terminal on insert)
|
| 克隆方法: | 限制性内切酶 |
| 5’测序引物 | f.hsyn |
| 3’测序引物 | r.WPRE |
|
文献引用方法 |
|
| 材料和方法部分: |
pAAV_hsyn_NES-his-CaMPARI2-WPRE-SV40 was a gift from Eric Schreiter (Addgene plasmid # 101060 ; http://n2t.net/addgene:101060 ; RRID:Addgene_101060)
For viral preps, please replace (Addgene plasmid # 101060) in the above sentence with: (Addgene viral prep # 101060-AAV1)
|
| 参考文献部分: | Improved methods for marking active neuron populations. Moeyaert B, Holt G, Madangopal R, Perez-Alvarez A, Fearey BC, Trojanowski NF, Ledderose J, Zolnik TA, Das A, Patel D, Brown TA, Sachdev RNS, Eickholt BJ, Larkum ME, Turrigiano GG, Dana H, Gee CE, Oertner TG, Hope BT, Schreiter ER. Nat Commun. 2018 Oct 25;9(1):4440. doi: 10.1038/s41467-018-06935-2. 10.1038/s41467-018-06935-2 PubMed 30361563 |
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
载体简介
载体序列
LOCUS pAAV_hsyn_NES-his-CaMPARI2-WPRE-SV40 5567 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION AAV vector expressing CaMPARI2 (Kd = 200nM), a photoconvertible
fluorescent protein-based calcium integrator, in neurons.
ACCESSION .
VERSION .
KEYWORDS pAAV_hsyn_NES-his-CaMPARI2-WPRE-SV40
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5567)
AUTHORS Moeyaert B, Holt G, Madangopal R, Perez-Alvarez A, Fearey BC,
Trojanowski NF, Ledderose J, Zolnik TA, Das A, Patel D, Brown TA,
Sachdev RNS, Eickholt BJ, Larkum ME, Turrigiano GG, Dana H, Gee CE,
Oertner TG, Hope BT, Schreiter ER
TITLE Improved methods for marking active neuron populations.
JOURNAL Nat Commun. 2018 Oct 25;9(1):4440. doi: 10.1038/s41467-018-06935-2.
PUBMED 30361563
REFERENCE 2 (bases 1 to 5567)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..5567
/organism="synthetic DNA construct"
/mol_type="other DNA"
promoter 197..644
/label=hSyn promoter
/note="human synapsin I promoter; confers neuron-specific
expression (Kugler et al., 2003)"
CDS 741..758
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
CDS 765..1205
/codon_start=1
/product="calcium-binding protein"
/label=calmodulin
/translation="DQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMRSLGQNPTEA
ELQDMINEVDADGDGTIDFPEFLTMMARKMKDTDSEEEIREAFRVFDKDGNGYISAAEL
RHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVVMMTAK"
CDS 2001..2024
/codon_start=1
/product="FLAG(R) epitope tag, followed by an enterokinase
cleavage site"
/label=FLAG
/translation="DYKDDDDK"
CDS 2043..2069
/codon_start=1
/product="HA (human influenza hemagglutinin) epitope tag"
/label=HA
/translation="YPYDVPDYA"
CDS 2088..2117
/codon_start=1
/product="Myc (human c-Myc proto-oncogene) epitope tag"
/label=Myc
/translation="EQKLISEEDL"
misc_feature 2139..2727
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
primer_bind complement(2192..2212)
/label=WPRE-R
/note="WPRE, reverse primer"
CDS complement(2610..2621)
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
primer_bind complement(2730..2746)
/label=KS primer
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(2731..2747)
/label=pBluescriptKS
/note="For pBluescript vector"
polyA_signal 2776..2897
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(2787..2806)
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind 2841..2860
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind complement(3105..3122)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(3276..3864)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(3356..3375)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(4035..4895)
/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 4658..4677
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(4896..5000)
/gene="bla"
/label=AmpR promoter
rep_origin complement(5026..5481)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(5163..5184)
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind 5375..5394
/label=F1ori-R
/note="F1 origin, reverse primer"
repeat_region 5553..115
/label=AAV2 ITR
ORIGIN
1 ctcactgagg ccgcccgggc aaagcccggg cgtcgggcga cctttggtcg cccggcctca
61 gtgagcgagc gagcgcgcag agagggagtg gccaactcca tcactagggg ttccttgtag
121 ttaatgatta acccgccatg ctacttatct acgtagccat gctctaggaa gatctctgca
181 gagggccctg cgtatgagtg caagtgggtt ttaggaccag gatgaggcgg ggtgggggtg
241 cctacctgac gaccgacccc gacccactgg acaagcaccc aacccccatt ccccaaattg
301 cgcatcccct atcagagagg gggaggggaa acaggatgcg gcgaggcgcg tgcgcactgc
361 cagcttcagc accgcggaca gtgccttcgc ccccgcctgg cggcgcgcgc caccgccgcc
421 tcagcactga aggcgcgctg acgtcactcg ccggtccccc gcaaactccc cttcccggcc
481 accttggtcg cgtccgcgcc gccgccggcc cagccggacc gcaccacgcg aggcgcgaga
541 taggggggca cgggcgcgac catctgcgct gcggcgccgg cgactcagcg ctgcctcagt
601 ctgcggtggg cagcggagga gtcgtgtcgt gcctgagagc gcagtcgaat tcaagctgct
661 agcaaggatc cgccaccatg ctgcagaacg agcttgctct taagttggct ggacttgata
721 ttaacaagac tggaggttct catcatcacc accaccatgg atccgaccaa ctgactgaag
781 agcagatcgc agaatttaaa gaggctttct ccctatttga caaggacggg gatgggacaa
841 taacaaccaa ggagctgggg acggtgatgc ggtctctggg gcagaacccc acagaagcag
901 agctgcagga catgatcaat gaagtagatg ccgacggtga cggcacaatc gacttccctg
961 agttcctgac aatgatggca agaaaaatga aagacacaga cagtgaagaa gaaattagag
1021 aagcgttccg tgtgtttgat aaggatggca atggctacat cagtgcagca gagcttcgcc
1081 acgtgatgac aaaccttgga gagaagttaa cagatgaaga ggttgatgaa atgatcaggg
1141 aagcagacat cgatggggat ggtcaggtaa actacgaaga gtttgtagtg atgatgacag
1201 cgaagctcga gtgcgagaaa atctatgtgc gtgatggagt gctgacgggt gatattcata
1261 tgatcttgtt gcttgaagga aatgcccatt accgatgtga cttcagaact acttacaaag
1321 ctaaggagaa gggtgtcaag ttaccaggcg tgcactatgt ggaccacacc attgagattt
1381 taagccatga caaagattac aacaaggtta agatctatga gtatgctgtt gctcattctg
1441 gattgcctga caatgccaga cgaggcggta ccggcggatc catggtgagt gcgattaagc
1501 cagacatgaa gatcaaactg cgtatggaag gcaacgtaaa cgggcaccac tttgtgatcg
1561 acggagatgg tacaggcaag ccttatgagg gaaaacagac catggatctt gaagtcaaag
1621 agggcggacc tctgcctttt gcctttgata tcctgaccac tgcattccat tacggcaaca
1681 gggtattcgt gaaatatcca gacaacatac aagactattt taagcagtcg tttcctaagg
1741 ggtattcgtg ggaacgaagc atgactttcg aagacggggg catttgctat gccagaagcg
1801 acataacaat ggaaggggac actttctata ataaagttcg attttatggt accaactttc
1861 ccgccaatgg tccagttatg cagaagaaga cgctgaaatg gatgccgagc tggacgcgtt
1921 catcacgtcg taagttcaat aagacaggtc acgcactgag agctataggt cggctgagct
1981 caggaggttc tggaggttct gattacaagg acgacgatga caagggaggt tctggaggtt
2041 cttacccata cgacgtacca gattacgctg gaggttctgg aggttctgag cagaagctga
2101 tctcagagga ggacctgcgt acgtaaaagc ttatcgataa tcaacctctg gattacaaaa
2161 tttgtgaaag attgactggt attcttaact atgttgctcc ttttacgcta tgtggatacg
2221 ctgctttaat gcctttgtat catgctattg cttcccgtat ggctttcatt ttctcctcct
2281 tgtataaatc ctggttgctg tctctttatg aggagttgtg gcccgttgtc aggcaacgtg
2341 gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg ttggggcatt gccaccacct
2401 gtcagctcct ttccgggact ttcgctttcc ccctccctat tgccacggcg gaactcatcg
2461 ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg
2521 tgttgtcggg gaaatcatcg tcctttcctt ggctgctcgc ctgtgttgcc acctggattc
2581 tgcgcgggac gtccttctgc tacgtccctt cggccctcaa tccagcggac cttccttccc
2641 gcggcctgct gccggctctg cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc
2701 ggatctccct ttgggccgcc tccccgcatc gataccgtcg acctcgaccc gggcggccgc
2761 ttcgagcaga catgataaga tacattgatg agtttggaca aaccacaact agaatgcagt
2821 gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa
2881 gctgcaataa acaagttaac aacaacaatt gcattcattt tatgtttcag gttcaggggg
2941 agatgtggga ggttttttaa agcaagtaaa acctctacaa atgtggtaaa atcgataagg
3001 atcttcctag agcatggcta cgtagataag tagcatggcg ggttaatcat taactacaag
3061 gaacccctag tgatggagtt ggccactccc tctctgcgcg ctcgctcgct cactgactcg
3121 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
3181 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
3241 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
3301 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
3361 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
3421 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
3481 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
3541 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
3601 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
3661 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
3721 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
3781 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
3841 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
3901 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
3961 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
4021 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
4081 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
4141 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
4201 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
4261 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
4321 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
4381 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
4441 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
4501 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
4561 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
4621 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
4681 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
4741 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
4801 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
4861 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
4921 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
4981 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctaaatt gtaagcgtta
5041 atattttgtt aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg
5101 ccgaaatcgg caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg
5161 ttccagtttg gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa
5221 aaaccgtcta tcagggcgat ggcccactac gtgaaccatc accctaatca agttttttgg
5281 ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt
5341 gacggggaaa gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg
5401 ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta
5461 atgcgccgct acagggcgcg tcccattcgc cattcaggct gcgcaactgt tgggaagggc
5521 gatcggtgcg ggcctcttcg ctattacgcc agctgcgcgc tcgctcg
//
