| 质粒名称: | pcDNA3-N-Flag-NLRP3 91-710 |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 75140 |
| 存储实验室: | Bruce Beutler |
|
载体骨架基本信息 |
|
| 载体名称: | pcDNA3 |
| 载体出品公司: | Clontech |
| 空载体大小: | 5446bp |
| 完整质粒大小: | 7321bp |
| 载体类型: | 哺乳细胞表达 |
| 筛选标记: | 新霉素（G418） |
| 载体抗性: | 氨苄霉素 |
| 生长温度: | 37 ℃ |
| 宿主菌: | DH5a |
| 拷贝数: | 未知 |
|
插入基因信息 |
|
| 插入基因名称: | NLR family, pyrin domain containing 3 / Nlrp3 |
| 物种来源: | M. musculus (mouse) |
| 插入大小: | 1875 bp |
| 基因突变: | fragment encoding amino acids 91-710 |
| GenBank编号 | NM_145827 |
| 启动子: | CMV |
| 标签/融合蛋白: | Flag (N terminal on insert) |
| 克隆方法: | 未知 |
| 5’测序引物 | CMV-F |
|
文献引用方法 |
|
| 材料和方法部分: |
pcDNA3-N-Flag-NLRP3 91-710 was a gift from Bruce Beutler (Addgene plasmid # 75140 ; http://n2t.net/addgene:75140 ; RRID:Addgene_75140)
|
| 参考文献部分: | NLRP3 activation and mitosis are mutually exclusive events coordinated by NEK7, a new inflammasome component. Shi H, Wang Y, Li X, Zhan X, Tang M, Fina M, Su L, Pratt D, Bu CH, Hildebrand S, Lyon S, Scott L, Quan J, Sun Q, Russell J, Arnett S, Jurek P, Chen D, Kravchenko VV, Mathison JC, Moresco EM, Monson NL, Ulevitch RJ, Beutler B. Nat Immunol. 2016 Mar;17(3):250-8. doi: 10.1038/ni.3333. Epub 2015 Dec 7. 10.1038/ni.3333 PubMed 26642356 |
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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 pcDNA3-N-Flag-NLRP3 91-710 7311 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION Expresses N-terminal Flag-tagged amino acids 91-710 of NLRP3 (the
nucleotide-binding domain) in mammalian cells.
ACCESSION .
VERSION .
KEYWORDS pcDNA3-N-Flag-NLRP3 91-710
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7311)
AUTHORS Shi H, Wang Y, Li X, Zhan X, Tang M, Fina M, Su L, Pratt D, Bu CH,
Hildebrand S, Lyon S, Scott L, Quan J, Sun Q, Russell J, Arnett S,
Jurek P, Chen D, Kravchenko VV, Mathison JC, Moresco EM, Monson NL,
Ulevitch RJ, Beutler B
TITLE NLRP3 activation and mitosis are mutually exclusive events
coordinated by NEK7, a new inflammasome component.
JOURNAL Nat Immunol. 2016 Mar;17(3):250-8. doi: 10.1038/ni.3333. Epub 2015
Dec 7.
PUBMED 26642356
REFERENCE 2 (bases 1 to 7311)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..7311
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind complement(44..63)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
enhancer 235..614
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 615..818
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 769..789
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
primer_bind 863..882
/label=T7
/note="T7 promoter, forward primer"
promoter 863..881
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
CDS 922..945
/codon_start=1
/product="FLAG(R) epitope tag, followed by an enterokinase
cleavage site"
/label=FLAG
/translation="DYKDDDDK"
promoter complement(2863..2881)
/label=SP6 promoter
/note="promoter for bacteriophage SP6 RNA polymerase"
primer_bind complement(2864..2881)
/label=SP6
/note="SP6 promoter, forward primer"
primer_bind complement(2901..2918)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 2907..3131
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin 3177..3605
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(3264..3283)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 3474..3495
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(3614..3634)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
promoter 3619..3949
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 3800..3935
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 3862..3881
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
CDS 4016..4810
/codon_start=1
/gene="aph(3')-II (or nptII)"
/product="aminoglycoside phosphotransferase from Tn5"
/label=NeoR/KanR
/note="confers resistance to neomycin, kanamycin, and G418
(Geneticin(R))"
/translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
primer_bind complement(4070..4089)
/label=Neo-R
/note="Neomycin resistance gene, reverse primer"
primer_bind 4680..4699
/label=Neo-F
/note="Neomycin resistance gene, forward primer"
polyA_signal 4984..5105
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(5021..5040)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 5075..5094
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(5154..5170)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(5154..5170)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(5167..5189)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 5178..5194
/label=lac operator
/bound_moiety="lac repressor encoded by lacI"
/note="The lac repressor binds to the lac operator to
inhibit transcription in E. coli. This inhibition can be
relieved by adding lactose or
isopropyl-beta-D-thiogalactopyranoside (IPTG)."
promoter complement(5202..5232)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 5247..5268
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(5385..5402)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(5556..6144)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(5636..5655)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(6315..7175)
/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 6938..6957
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(7176..7280)
/gene="bla"
/label=AmpR promoter
ORIGIN
1 gacggatcgg gagatctccc gatcccctat ggtcgactct cagtacaatc tgctctgatg
61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
421 attgacgtca atgggtggac tatttacggt aaactgccca cttggcagta catcaagtgt
481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttggtacc
901 gagctcggat cgcccaccat ggactacaaa gacgatgacg ataaaatcga tatcgaattc
961 atgagtggaa tgacacgtgt acatcacatt cctctatggt atgccaggag gacagccttg
1021 aagaagagtg gatgggtttg ctgggatatc tctcccgcat ctccatttgt aaaaagaaga
1081 aagattactg taagatgtac agacgacatg tgagaagcag gttctactct atcaaggaca
1141 ggaacgcgcg tctaggtgag agtgtggacc tcaacagtcg ctacacgcag ctccaactgg
1201 tcaaggagca tccaagcaag caggagcggg agcatgaact cctgaccatc ggccggacta
1261 aaatgcggga cagccccatg agttccctta agctggagct gctgtttgag cccgaggacg
1321 ggcactcgga gcctgtgcac acagtggtgt tccagggagc agcaggcatc gggaaaacca
1381 tcctagccag gaagattatg ttggactggg cactgggaaa gctcttcaaa gacaaatttg
1441 actatttgtt ctttatccac tgccgagagg tgagcctcag gacgccaagg agtctagcag
1501 acctgattgt cagctgctgg cctgacccaa acccaccagt gtgcaagatc ctgcgcaagc
1561 cttccaggat cctcttcctc atggatggct ttgatgagct acaaggggcc tttgacgagc
1621 acattgggga ggtctgcaca gactggcaaa aggctgtgcg gggagacatt ctgctaagca
1681 gcctcatccg aaagaaactg ctgcccaagg cctctctgct cataacgacg aggccggtag
1741 ccttggagaa actgcagcat ctcctggacc acccccgcca tgtggagatc ctaggtttct
1801 ctgaggccaa aaggaaggag tatttcttta agtatttctc caacgagctg caggcccggg
1861 aggccttcag gctgatccaa gagaatgagg tcctctttac catgtgcttc atccccctgg
1921 tctgctggat tgtgtgcacg gggctaaagc aacagatgga gaccgggaag agcctggccc
1981 agacctccaa gaccactacg gccgtctacg tcttcttcct ttccagcctg ctgcaatccc
2041 gggggggcat tgaggagcat ctcttctctg actacctaca ggggctctgt tcactggctg
2101 cggatggaat ttggaaccag aaaatcctat ttgaggagtg tgatctgcgg aagcacggcc
2161 tgcagaagac tgacgtctcc gctttcctga ggatgaacgt gttccagaag gaagtggact
2221 gcgagagatt ctacagcttc agccacatga ctttccagga gttcttcgct gctatgtact
2281 atttgctgga agaggaggca gagggggaga ccgtgaggaa aggaccagga ggttgttcag
2341 atcttctgaa ccgagacgtg aaggtcctac tagaaaatta cggcaagttt gaaaaaggct
2401 atctgatttt tgttgtccga ttcctctttg gccttgtaaa ccaggagaga acctcttatt
2461 tggagaagaa actaagttgc aagatctctc agcaagtcag actggaacta ctgaagtgga
2521 ttgaagtgaa agccaaggcc aagaagctgc agtggcagcc cagccaactg gaactgttct
2581 actgcctgta cgagatgcag gaggaagact ttgtgcagag tgccatggac cactttccca
2641 aaattgagat caacctctct accagaatgg accacgtggt ttcctccttt tgtattaaga
2701 actgtcatag ggtcaaaacg ctttccctgg gtttttttca caactcgccc aaggaggaag
2761 aagaagagag gagaggaggt cgacccttgg accaggttca gtgtgttttc ccagacactc
2821 atgttgcctg tttagctcga gcatgcatct agagggccct attctatagt gtcacctaaa
2881 tgctagagct cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg
2941 cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata
3001 aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt
3061 ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt
3121 gggctctatg gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc
3181 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
3241 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
3301 cgccggcttt ccccgtcaag ctctaaatcg gggcatccct ttagggttcc gatttagtgc
3361 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
3421 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
3481 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
3541 gattttgggg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
3601 gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccaggc
3661 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
3721 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
3781 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
3841 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct
3901 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg
3961 agcttgtata tccattttcg gatctgatca agagacagga tgaggatcgt ttcgcatgat
4021 tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta
4081 tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca
4141 ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcagga
4201 cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga
4261 cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct
4321 cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg
4381 gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga
4441 gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca
4501 tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga
4561 ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg
4621 cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc
4681 gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt
4741 gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga
4801 gttcttctga gcgggactct ggggttcgaa atgaccgacc aagcgacgcc caacctgcca
4861 tcacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc
4921 cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac
4981 cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
5041 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
5101 tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc atggtcatag
5161 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
5221 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
5281 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
5341 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
5401 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
5461 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
5521 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
5581 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
5641 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
5701 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca atgctcacgc
5761 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
5821 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
5881 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
5941 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca
6001 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
6061 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
6121 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
6181 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
6241 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
6301 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
6361 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
6421 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
6481 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
6541 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
6601 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
6661 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
6721 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
6781 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
6841 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
6901 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
6961 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
7021 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
7081 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
7141 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
7201 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
7261 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt c
//
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