| 质粒名称: | pAAV-EF1a-DIO-hM4D(Gi)-mCherry |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 50461 |
| 存储实验室: | Bryan Roth |
|
载体骨架基本信息 |
|
| 载体名称: | pAAV |
| 载体出品公司: | Clontech |
| 空载体大小: | 4818bp |
| 完整质粒大小: | 7772bp |
| 载体类型: | AAV |
| 载体抗性: | 氨苄霉素 |
| 生长温度: | 37 ℃ |
| 宿主菌: | Stbl3 |
| 拷贝数: | 低拷贝 |
|
插入基因信息 |
|
| 插入基因名称: | hM4D(Gi)-mCherry |
| 物种来源: | H. sapiens (human) |
| 插入大小: | 2175bp |
| 启动子: | EF1a |
| 标签/融合蛋白: |
HA (N terminal on insert)
mCherry (C terminal on insert)
|
| 克隆方法: | 限制性内切酶 |
| 5’克隆位点: | Asc I (not destroyed) |
| 3’克隆位点: | Nhe I (not destroyed) |
| 5’测序引物 | TTTTTGAGTTTGGATCTTGG |
| 3’测序引物 | GCATTAAAGCAGCGTATCCACATAGC |
|
文献引用方法 |
|
| 材料和方法部分: | pAAV-EF1a-DIO-hM4D(Gi)-mCherry was a gift from Bryan Roth (Addgene plasmid # 50461 ; http://n2t.net/addgene:50461 ; RRID:Addgene_50461) |
买家导航
pAAV-EF1a-DIO-hM4D(Gi)-mCherry载体质粒基本信息
pAAV-EF1a-DIO-hM4D(Gi)-mCherry质粒图谱载体图谱和pAAV-EF1a-DIO-hM4D(Gi)-mCherry载体序列质粒序列多克隆位点信息
-mCherry质粒图谱.png)
pAAV-EF1a-DIO-hM4D(Gi)-mCherry质粒载体简介
pAAV-EF1a-DIO-hM4D(Gi)-mCherry质粒序列载体序列
LOCUS pAAV-EF1a-DIO-hM4D(Gi)-mCherry 7762 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION Double floxed Gi-coupled hM4D DREADD fused with mCherry under the
control of EF1a promoter .
ACCESSION .
VERSION .
KEYWORDS pAAV-EF1a-DIO-hM4D(Gi)-mCherry
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7762)
AUTHORS Bryan Roth
TITLE Roth lab DREADDs
JOURNAL Unpublished
REFERENCE 2 (bases 1 to 7762)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..7762
/organism="synthetic DNA construct"
/mol_type="other DNA"
regulatory 599..608
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 605..1315
/codon_start=1
/product="monomeric derivative of DsRed fluorescent protein
(Shaner et al., 2004)"
/label=mCherry
/note="mammalian codon-optimized"
/translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
EGRHSTGGMDELYK"
primer_bind complement(750..768)
/label=mCherry-R
/note="mCherry, reverse primer"
primer_bind 1018..1037
/label=mCherry-F
/note="mCherry, forward primer"
protein_bind complement(1324..1357)
/label=loxP
/bound_moiety="Cre recombinase"
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (GCATACAT)."
protein_bind complement(1408..1441)
/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(1472..2650)
/label=EF-1-alpha promoter
/note="strong constitutive promoter for human elongation
factor EF-1-alpha"
intron 1481..2419
/label=EF-1-alpha intron A
/note="intron upstream of the start codon of human
EF-1-alpha"
primer_bind complement(1504..1524)
/label=EF1a-F
/note="Human elongation factor-1a promoter, forward primer"
repeat_region 2751..2880
/label=AAV2 ITR (alternate)
/note="Functional equivalent of wild-type AAV2 ITR"
rep_origin complement(2942..3530)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(3022..3041)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(3701..4561)
/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 4324..4343
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(4562..4666)
/gene="bla"
/label=AmpR promoter
primer_bind 4734..4752
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
primer_bind complement(4790..4812)
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind 4912..4931
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
rep_origin complement(4948..5403)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(5085..5106)
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind 5297..5316
/label=F1ori-R
/note="F1 origin, reverse primer"
repeat_region 5478..5618
/label=AAV2 ITR
/note="inverted terminal repeat of adeno-associated virus
serotype 2"
repeat_region 5489..5618
/label=AAV2 ITR
polyA_signal 5658..6134
/label=hGH poly(A) signal
/note="human growth hormone polyadenylation signal"
primer_bind 5916..5935
/label=hGH-PA-R
/note="Human growth hormone terminator, reverse primer"
misc_feature 6166..6754
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
CDS 6272..6283
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
primer_bind 6681..6701
/label=WPRE-R
/note="WPRE, reverse primer"
protein_bind 6779..6812
/label=loxP
/bound_moiety="Cre recombinase"
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (GCATACAT)."
protein_bind 6863..6896
/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 6903..6912
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
ORIGIN
1 ccccgtggct gataaggaca cttccaatga gtccagctca ggcagtgcca cccagaacac
61 caaggaacgc ccagccacag agctgtccac cacagaggcc accacgcccg ccatgcccgc
121 ccctcccctg cagccgcggg ccctcaaccc agcctccaga tggtccaaga tccagattgt
181 gacgaagcag acaggcaatg agtgtgtgac agccattgag attgtgcctg ccacgccggc
241 tggcatgcgc cctgcggcca acgtggcccg caagttcgcc agcatcgctc gcaaccaggt
301 gcgcaagaag cggcagatgg cggcccggga gcgcaaagtg acacgaacga tctttgccat
361 tctgctggcc ttcatcctca cctggacgcc ctacaacgtc atggtcctgg tgaacacctt
421 ctgccagagc tgcatccctg acacggtgtg gtccattggc tactggctct gctacgtcaa
481 cagcaccatc aaccctgcct gctatgctct gtgcaacgcc acctttaaaa agaccttccg
541 gcacctgctg ctgtgccagt atcggaacat cggcactgcc aggcgggatc caccggtcgc
601 caccatggtg agcaagggcg aggaggataa catggccatc atcaaggagt tcatgcgctt
661 caaggtgcac atggagggct ccgtgaacgg ccacgagttc gagatcgagg gcgagggcga
721 gggccgcccc tacgagggca cccagaccgc caagctgaag gtgaccaagg gtggccccct
781 gcccttcgcc tgggacatcc tgtcccctca gttcatgtac ggctccaagg cctacgtgaa
841 gcaccccgcc gacatccccg actacttgaa gctgtccttc cccgagggct tcaagtggga
901 gcgcgtgatg aacttcgagg acggcggcgt ggtgaccgtg acccaggact cctccctgca
961 ggacggcgag ttcatctaca aggtgaagct gcgcggcacc aacttcccct ccgacggccc
1021 cgtaatgcag aagaagacca tgggctggga ggcctcctcc gagcggatgt accccgagga
1081 cggcgccctg aagggcgaga tcaagcagag gctgaagctg aaggacggcg gccactacga
1141 cgctgaggtc aagaccacct acaaggccaa gaagcccgtg cagctgcccg gcgcctacaa
1201 cgtcaacatc aagttggaca tcacctccca caacgaggac tacaccatcg tggaacagta
1261 cgaacgcgcc gagggccgcc actccaccgg cggcatggac gagctgtaca agtaaggcgc
1321 gccataactt cgtataatgt atgctatacg aagttattaa gaggtttcat attgctaata
1381 gcagctacaa tccagctacc attctgcata acttcgtata aagtatccta tacgaagtta
1441 ttccggagtc gactctagag gatccggtac ctcacgacac ctgaaatgga agaaaaaaac
1501 tttgaaccac tgtctgaggc ttgagaatga accaagatcc aaactcaaaa agggcaaatt
1561 ccaaggagaa ttacatcaag tgccaagctg gcctaacttc agtctccacc cactcagtgt
1621 ggggaaactc catcgcataa aacccctccc cccaacctaa agacgacgta ctccaaaagc
1681 tcgagaacta atcgaggtgc ctggacggcg cccggtactc cgtggagtca catgaagcga
1741 cggctgagga cggaaaggcc cttttccttt gtgtgggtga ctcacccgcc cgctctcccg
1801 agcgccgcgt cctccatttt gagctccctg cagcagggcc gggaagcggc catctttccg
1861 ctcacgcaac tggtgccgac cgggccagcc ttgccgccca gggcggggcg atacacggcg
1921 gcgcgaggcc aggcaccaga gcaggccggc cagcttgaga ctacccccgt ccgattctcg
1981 gtggccgcgc tcgcaggccc cgcctcgccg aacatgtgcg ctgggacgca cgggccccgt
2041 cgccgcccgc ggccccaaaa accgaaatac cagtgtgcag atcttggccc gcatttacaa
2101 gactatcttg ccagaaaaaa agcgtcgcag caggtcatca aaaattttaa atggctagag
2161 acttatcgaa agcagcgaga caggcgcgaa ggtgccacca gattcgcacg cggcggcccc
2221 agcgcccagg ccaggcctca actcaagcac gaggcgaagg ggctccttaa gcgcaaggcc
2281 tcgaactctc ccacccactt ccaacccgaa gctcgggatc aagaatcacg tactgcagcc
2341 aggtggaagt aattcaaggc acgcaagggc cataacccgt aaagaggcca ggcccgcggg
2401 aaccacacac ggcacttacc tgtgttctgg cggcaaaccc gttgcgaaaa agaacgttca
2461 cggcgactac tgcacttata tacggttctc ccccaccctc gggaaaaagg cggagccagt
2521 acacgacatc actttcccag tttaccccgc gccaccttct ctaggcaccg gttcaattgc
2581 cgacccctcc ccccaacttc tcggggactg tgggcgatgt gcgctctgcc cactgacggg
2641 caccggagcc aattcccact cctttcaaga cctagaaggt ccattagctg caaagattcc
2701 tctctgttta aaactttatc catctttgca aagcttacgc gtgcggccgc aggaacccct
2761 agtgatggag ttggccactc cctctctgcg cgctcgctcg ctcactgagg ccgggcgacc
2821 aaaggtcgcc cgacgcccgg gcggcctcag tgagcgagcg agcgcgcagc tgcctgcagg
2881 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
2941 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
3001 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
3061 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
3121 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
3181 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
3241 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
3301 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
3361 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
3421 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
3481 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
3541 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
3601 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
3661 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
3721 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
3781 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
3841 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
3901 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
3961 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
4021 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
4081 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
4141 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
4201 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
4261 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
4321 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
4381 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
4441 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
4501 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
4561 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
4621 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
4681 aagtgccacc tgacgtctaa gaaaccatta ttatcatgac attaacctat aaaaataggc
4741 gtatcacgag gccctttcgt ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca
4801 tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc
4861 gtcagggcgc gtcagcgggt gttggcgggt gtcggggctg gcttaactat gcggcatcag
4921 agcagattgt actgagagtg caccataaaa ttgtaaacgt taatattttg ttaaaattcg
4981 cgttaaattt ttgttaaatc agctcatttt ttaaccaata gaccgaaatc ggcaaaatcc
5041 cttataaatc aaaagaatag cccgagatag agttgagtgt tgttccagtt tggaacaaga
5101 gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg
5161 atggcccact acgtgaacca tcacccaaat caagtttttt ggggtcgagg tgccgtaaag
5221 cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga
5281 acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctaaggcg ctggcaagtg
5341 tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg
5401 cgtactatgg ttgctttgac gtatgcggtg tgaaataccg cacagatgcg taaggagaaa
5461 ataccgcatc aggcgcccct gcaggcagct gcgcgctcgc tcgctcactg aggccgcccg
5521 ggcaaagccc gggcgtcggg cgacctttgg tcgcccggcc tcagtgagcg agcgagcgcg
5581 cagagaggga gtggccaact ccatcactag gggttcctgc ggccgctcgg tccgcacgtg
5641 gttacctaca aaatcagaag gacagggaag ggagcagtgg ttcacgcctg taatcccagc
5701 aatttgggag gccaaggtgg gtagatcacc tgagattagg agttggagac cagcctggcc
5761 aatatggtga aaccccgtct ctaccaaaaa aacaaaaatt agctgagcct ggtcatgcat
5821 gcctggaatc ccaacaactc gggaggctga ggcaggagaa tcgcttgaac ccaggaggcg
5881 gagattgcag tgagccaaga ttgtgccact gcactccagc ttggttccca atagaccccg
5941 caggccctac aggttgtctt cccaacttgc cccttgctcc ataccacccc cctccacccc
6001 ataatattat agaaggacac ctagtcagac aaaatgatgc aacttaattt tattaggaca
6061 aggctggtgg gcactggagt ggcaacttcc agggccagga gaggcactgg ggaggggtca
6121 cagggatgcc acccgtagat ctctcgagca gcgctcggta tcgatgcggg gaggcggccc
6181 aaagggagat ccgactcgtc tgagggcgaa ggcgaagacg cggaagaggc cgcagagccg
6241 gcagcaggcc gcgggaagga aggtccgctg gattgagggc cgaagggacg tagcagaagg
6301 acgtcccgcg cagaatccag gtggcaacat aggcgagcag ccaaggaaag gacgatgatt
6361 tccccgacaa caccacggaa ttgtcagtgc ccaacagccg agcccctgtc cagcagcggg
6421 caaggcaggc ggcgatgagt tccgccgtgg caatagggag ggggaaagcg aaagtcccgg
6481 aaaggagctg acaggtggtg gcaatgcccc aaccagtggg ggttgcgtca gcaaacacag
6541 tgcacaccac gccacgttgc ctgacaacgg gccacaactc ctcataaaga gacagcaacc
6601 aggatttata caaggaggag aaaatgaaag ccatacggga agcaatagca tgatacaaag
6661 gcattaaagc agcgtatcca catagcgtaa aaggagcaac atagttaaga ataccagtca
6721 atctttcaca aattttgtaa tccagaggtt gattatcgat aagcttgata tcgaattcat
6781 aacttcgtat agcatacatt atacgaagtt attctttgca ccattctaaa gaataacagt
6841 gataatttct gggttaaggc aaataacttc gtataggata ctttatacga agttatgcta
6901 gcgccaccat ggccaacttc acacctgtca atggcagctc gggcaatcag tccgtgcgcc
6961 tggtcacgtc atcatcccac aatcgctatg agacggtgga aatggtcttc attgccacag
7021 tgacaggctc cctgagcctg gtgactgtcg tgggcaacat cctggtgatg ctgtccatca
7081 aggtcaacag gcagctgcag acagtcaaca actacttcct cttcagcctg gcgtgtgctg
7141 atctcatcat aggcgccttc tccatgaacc tctacaccgt gtacatcatc aagggctact
7201 ggcccctggg cgccgtggtc tgcgacctgt ggctggccct ggactgcgtg gtgagcaacg
7261 cctccgtcat gaaccttctc atcatcagct ttgaccgcta cttctgcgtc accaagcctc
7321 tcacctaccc tgcccggcgc accaccaaga tggcaggcct catgattgct gctgcctggg
7381 tactgtcctt cgtgctctgg gcgcctgcca tcttgttctg gcagtttgtg gtgggtaagc
7441 ggacggtgcc cgacaaccag tgcttcatcc agttcctgtc caacccagca gtgacctttg
7501 gcacagccat tgctggcttc tacctgcctg tggtcatcat gacggtgctg tacatccaca
7561 tctccctggc cagtcgcagc cgagtccaca agcaccggcc cgagggcccg aaggagaaga
7621 aagccaagac gctggccttc ctcaagagcc cactaatgaa gcagagcgtc aagaagcccc
7681 cgcccgggga ggccgcccgg gaggagctgc gcaatggcaa gctggaggag gcccccccgc
7741 cagcgctgcc accgccaccg cg
//
