pLVX-PTuner

价格:3500元

联系方式:I47-825O-882O

买家导航

载体基本信息

出品公司: Clontech
载体名称: pLVX-PTuner
质粒类型: 慢病毒载体;双顺反子载体
高拷贝/低拷贝: 高拷贝
克隆方法: 限制性内切酶,多克隆位点
启动子: CMV
载体大小: 8473 bp
5' 测序引物及序列: CMV Forward: CGCAAATGGGCGGTAGGCGTG
3' 测序引物及序列: --
载体标签: DD tag (N-端)
载体抗性: 氨苄青霉素
筛选标记: 嘌呤霉素(Puromycin)
克隆菌株: Stbl3
宿主细胞(系): 常规细胞系(293、CV-1、CHO等)
备注: 慢毒载体pLVX-PTuner载体表达N端DD标签融合蛋白;
DD标签是一个大小为12KD的FKBP (L106P)去稳定结构域,带有DD标签的目的蛋白将很快被降解;
向培养基中加入Shield1配体后,情况将发生逆转,细胞中目的蛋白的量则迅速提高。
产品目录号: 632173
稳定性: 稳表达
组成型/诱导型: 组成型
病毒/非病毒: 慢病毒

载体质粒图谱和多克隆位点信息

pLVX-PTuner载体图谱



pLVX-PTuner 多克隆位点

pLVX-PTuner 载体特征

载体简介

pLVX-PTuner载体描述

pLVX-PTuner is a bicistronic, HIV-1-based, lentiviral expression vector that allows you to precisely regulate the amount of your protein of interest in virtually any mammalian cell type, including primary cells. The vector encodes a 12 kDa, FKBP (L106P) destabilization domain (DD; 1) that is expressed as an N-terminal tag on your protein of interest, causing rapid degradation of the fusion protein. Degradation of the DD-tagged protein can be prevented by the addition of Shield1 stabilizing ligand to the medium. Shield1 is a membrane permeable molecule that binds to the DD tag, 'shielding' the fusion protein from proteasomal degradation.
pLVX-PTuner allows the simultaneous expression of your DD-tagged protein of interest and a puromycin resistance marker (Puror) from the same bicistronic mRNA transcript. Expression of this transcript is driven by the constitutively active human cytomegalovirus immediate early promoter (PCMV IE) located just upstream of the DD tag coding sequence. An encephalomyocarditis virus (EMCV) internal ribosome entry site (IRES), positioned between the MCS and Puror (above), facilitates cap-independent translation of Puror from an internal start site at the IRES/Puror junction (2). Because Puror is unaffected by the DD tag, puromycin resistance can be used as an indicator of transduction efficiency and a marker for selection.
pLVX-PTuner contains all of the viral processing elements necessary for the production of replication-incompetent lentivirus, as well as elements to improve viral titer, transgene expression, and overall vector function. The woodchuck hepatitis virus posttranscriptional regulatory element (WPRE) promotes RNA processing events and enhances nuclear export of viral and transgene RNA (3), leading to increased viral titers from packaging cells and enhanced expression of your gene of interest in target cells. In addition, the vector includes a Rev-response element (RRE), which further increases viral titers by enhancing the transport of unspliced viral RNA out of the nucleus (4). Finally, pLVX-PTuner also contains a central polypurine tract/central termination sequence element (cPPT/CTS). During target cell infection, this element creates a central DNA flap that increases nuclear import of the viral genome, resulting in improved vector integration and more efficient transduction (5). The vector also contains a pUC origin of replication and an E. coli ampicillin resistance gene (Ampr) for propagation and selection in bacteria.

pLVX-PTuner is available in the Lenti-X ProteoTuner Shield System N (Cat. No. 632173). The vector is designed to constitutively coexpress a DD-tagged protein of interest and puromycin resistance from PCMV IE when transduced into mammalian cells. In order to create your N-terminally tagged protein of interest, your gene of interest must be cloned into the MCS in the same reading frame as the DD tag sequence, and it must contain a stop codon at the end of its coding sequence. Before it can be transduced into target cells, the vector must be cotransfected into 293T cells with our Lenti-X HTX Packaging System (Cat. Nos. 631247 and 631249) and packaged into viral particles. This packaging system allows the safe production of high titer, infectious, replication-incompetent, VSV-G pseudotyped lentiviral particles that can infect a wide range of cell types, including non-dividing and primary cells (6).
When cells expressing a DD-tagged protein of interest are grown in medium containing Shield1, the ligand binds to the DD tag and protects the fusion protein from degradation. As a result, the protein quickly accumulates inside the cells in amounts that are directly proportional to the concentration of Shield1 in the medium. If the cells are subsequently grown in medium lacking Shield1, the DD tag is no longer stabilized, and the fusion protein is rapidly degraded. Because the effects of Shield1 are concentration-dependent and reversible, it is possible to fine-tune the amount of fusion protein present in the cells simply by adjusting the concentration of the ligand in the medium (1).

Propagation in E. coli
 Suitable host strains: Stellar Competent Cells.
 Selectable marker: plasmid confers resistance to ampicillin (100 μg/ml) in E. coli hosts.
 E. coli replication origin: pUC
 Copy number: high

Notes:
The vector sequence was compiled from information in the sequence databases, published literature, and other sources, together with partial sequences obtained by Clontech. This vector has not been completely sequenced.
The viral supernatants produced by this lentiviral vector could contain potentially hazardous recombinant virus. Due caution must be exercised in the production and handling of recombinant lentivirus. Appropriate NIH, regional, and institutional guidelines apply.

References
1. Banaszynski, L. et al. (2006) Cell 126(5):995–1004.
2. Jang, S. K. et al. (1988) J. Virol. 62(8):2636–2643.
3. Zufferey, R. et al. (1999) J. Virol. 73(4):2886–2892.
4. Cochrane, A. W. et al. (1990) Proc. Natl. Acad. Sci. USA 87(3):1198–1202.
5. Zennou, V. et al. (2000) Cell 101(2):173–185.
6. Wu, X. et al. (2000) Mol. Ther. 2(1):47–55.


载体序列

LOCUS       pLVX-PTuner   8472 bp    DNA   circular             8-NOV-2009
COMMENT     Created by Clontech Laboratories Inc. http://www.clontech.com
COMMENT     Cat. No. 632173,632174
FEATURES             Location/Qualifiers
     LTR             1..635
                     /label=5'\LTR
     misc_feature    685..822
                     /label=packaging\signal
     misc_feature    2028..2151
                     /label=cPPT/CTS\(DNA\flap)
     promoter        2185..2787
                     /label=CMV\IE\Promoter
     misc_feature    4394..4985
                     /label=WPRE
     rep_origin      complement(6293..6966)
                     /label=pUC\origin
     CDS             complement(7111..8107)
                     /label=Amp(R)
     misc_binding    1303..1536
                     /label=RRE
     splicing_signal 744..745
                     /label=SD
     splicing_signal 1883..1884
                     /label=SA
     misc_binding    636..653
                     /label=PBS
     LTR             5188..5824
                     /label=3'\LTR
     CDS             2805..3128
                     /label=DD
     misc_feature    3173..3780
                     /label=IRES
     CDS             3781..4380
                     /label=Puro(R)
BASE COUNT     2170 a      2109 c      2188 g      2005 t 
ORIGIN
        1 tggaagggct aattcactcc caaagaagac aagatatcct tgatctgtgg atctaccaca 
       61 cacaaggcta cttccctgat tagcagaact acacaccagg gccaggggtc agatatccac 
      121 tgacctttgg atggtgctac aagctagtac cagttgagcc agataaggta gaagaggcca 
      181 ataaaggaga gaacaccagc ttgttacacc ctgtgagcct gcatgggatg gatgacccgg 
      241 agagagaagt gttagagtgg aggtttgaca gccgcctagc atttcatcac gtggcccgag 
      301 agctgcatcc ggagtacttc aagaactgct gatatcgagc ttgctacaag ggactttccg 
      361 ctggggactt tccagggagg cgtggcctgg gcgggactgg ggagtggcga gccctcagat 
      421 cctgcatata agcagctgct ttttgcctgt actgggtctc tctggttaga ccagatctga 
      481 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct 
      541 tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc 
      601 agaccctttt agtcagtgtg gaaaatctct agcagtggcg cccgaacagg gacttgaaag 
      661 cgaaagggaa accagaggag ctctctcgac gcaggactcg gcttgctgaa gcgcgcacgg 
      721 caagaggcga ggggcggcga ctggtgagta cgccaaaaat tttgactagc ggaggctaga 
      781 aggagagaga tgggtgcgag agcgtcagta ttaagcgggg gagaattaga tcgcgatggg 
      841 aaaaaattcg gttaaggcca gggggaaaga aaaaatataa attaaaacat atagtatggg 
      901 caagcaggga gctagaacga ttcgcagtta atcctggcct gttagaaaca tcagaaggct 
      961 gtagacaaat actgggacag ctacaaccat cccttcagac aggatcagaa gaacttagat 
     1021 cattatataa tacagtagca accctctatt gtgtgcatca aaggatagag ataaaagaca 
     1081 ccaaggaagc tttagacaag atagaggaag agcaaaacaa aagtaagacc accgcacagc 
     1141 aagcggccgg ccgctgatct tcagacctgg aggaggagat atgagggaca attggagaag 
     1201 tgaattatat aaatataaag tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc 
     1261 aaagagaaga gtggtgcaga gagaaaaaag agcagtggga ataggagctt tgttccttgg 
     1321 gttcttggga gcagcaggaa gcactatggg cgcagcgtca atgacgctga cggtacaggc 
     1381 cagacaatta ttgtctggta tagtgcagca gcagaacaat ttgctgaggg ctattgaggc 
     1441 gcaacagcat ctgttgcaac tcacagtctg gggcatcaag cagctccagg caagaatcct 
     1501 ggctgtggaa agatacctaa aggatcaaca gctcctgggg atttggggtt gctctggaaa 
     1561 actcatttgc accactgctg tgccttggaa tgctagttgg agtaataaat ctctggaaca 
     1621 gatttggaat cacacgacct ggatggagtg ggacagagaa attaacaatt acacaagctt 
     1681 aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt 
     1741 ggaattagat aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt ggctgtggta 
     1801 tataaaatta ttcataatga tagtaggagg cttggtaggt ttaagaatag tttttgctgt 
     1861 actttctata gtgaatagag ttaggcaggg atattcacca ttatcgtttc agacccacct 
     1921 cccaaccccg aggggacccg acaggcccga aggaatagaa gaagaaggtg gagagagaga 
     1981 cagagacaga tccattcgat tagtgaacgg atctcgacgg tatcgccttt aaaagaaaag 
     2041 gggggattgg ggggtacagt gcaggggaaa gaatagtaga cataatagca acagacatac 
     2101 aaactaaaga attacaaaaa caaattacaa aaattcaaaa ttttcgggtt tattacaggg 
     2161 acagcagaga tccagtttat cgataagctt gggagttccg cgttacataa cttacggtaa 
     2221 atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg 
     2281 ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt 
     2341 aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg 
     2401 tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc 
     2461 ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc 
     2521 agtacatcaa tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca 
     2581 ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta 
     2641 acaactccgc cccattgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa 
     2701 gcagagctcg tttagtgaac cgtcagatcg cctggagacg ccatccacgc tgttttgacc 
     2761 tccatagaag acaccgactc tactagagga tctatttaaa taccatggga gtgcaggtgg 
     2821 aaaccatctc cccaggagac gggcgcacct tccccaagcg cggccagacc tgtgtggtgc 
     2881 actacaccgg gatgcttgaa gatggaaaga aagtcgattc ctcccgggac agaaacaagc 
     2941 cctttaagtt tatgctaggc aagcaggagg tgatccgagg ctgggaagaa ggggttgccc 
     3001 agatgagtgt gggtcagaga gccaaactga ctatatctcc agattatgcc tatggtgcca 
     3061 ctgggcaccc aggcatcatc ccaccacatg ccactctcgt cttcgatgtg gagcttctaa 
     3121 aaccggaaac cggtgaattc ctcgagacta gttctagagc ggccgcggat cccgcccctc 
     3181 tccctccccc ccccctaacg ttactggccg aagccgcttg gaataaggcc ggtgtgcgtt 
     3241 tgtctatatg ttattttcca ccatattgcc gtcttttggc aatgtgaggg cccggaaacc 
     3301 tggccctgtc ttcttgacga gcattcctag gggtctttcc cctctcgcca aaggaatgca 
     3361 aggtctgttg aatgtcgtga aggaagcagt tcctctggaa gcttcttgaa gacaaacaac 
     3421 gtctgtagcg accctttgca ggcagcggaa ccccccacct ggcgacaggt gcctctgcgg 
     3481 ccaaaagcca cgtgtataag atacacctgc aaaggcggca caaccccagt gccacgttgt 
     3541 gagttggata gttgtggaaa gagtcaaatg gctctcctca agcgtattca acaaggggct 
     3601 gaaggatgcc cagaaggtac cccattgtat gggatctgat ctggggcctc ggtgcacatg 
     3661 ctttacatgt gtttagtcga ggttaaaaaa acgtctaggc cccccgaacc acggggacgt 
     3721 ggttttcctt tgaaaaacac gatgataagc ttgccacaac ccacaaggag acgaccttcc 
     3781 atgaccgagt acaagcccac ggtgcgcctc gccacccgcg acgacgtccc ccgggccgta 
     3841 cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc gccacaccgt cgacccggac 
     3901 cgccacatcg agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac 
     3961 atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac cacgccggag 
     4021 agcgtcgaag cgggggcggt gttcgccgag atcggcccgc gcatggccga gttgagcggt 
     4081 tcccggctgg ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg gcccaaggag 
     4141 cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc accagggcaa gggtctgggc 
     4201 agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc cgccttcctg 
     4261 gagacctccg cgccccgcaa cctccccttc tacgagcggc tcggcttcac cgtcaccgcc 
     4321 gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc cggtgcctag 
     4381 acgcgtctgg aacaatcaac ctctggatta caaaatttgt gaaagattga ctggtattct 
     4441 taactatgtt gctcctttta cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc 
     4501 tattgcttcc cgtatggctt tcattttctc ctccttgtat aaatcctggt tgctgtctct 
     4561 ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga 
     4621 cgcaaccccc actggttggg gcattgccac cacctgtcag ctcctttccg ggactttcgc 
     4681 tttccccctc cctattgcca cggcggaact catcgccgcc tgccttgccc gctgctggac 
     4741 aggggctcgg ctgttgggca ctgacaattc cgtggtgttg tcggggaagc tgacgtcctt 
     4801 tccatggctg ctcgcctgtg ttgccacctg gattctgcgc gggacgtcct tctgctacgt 
     4861 cccttcggcc ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc 
     4921 tcttccgcgt cttcgccttc gccctcagac gagtcggatc tccctttggg ccgcctcccc 
     4981 gcctggaatt aattctgcag tcgagaccta gaaaaacatg gagcaatcac aagtagcaat 
     5041 acagcagcta ccaatgctga ttgtgcctgg ctagaagcac aagaggagga ggaggtgggt 
     5101 tttccagtca cacctcaggt acctttaaga ccaatgactt acaaggcagc tgtagatctt 
     5161 agccactttt taaaagaaaa gaggggactg gaagggctaa ttcactccca acgaagacaa 
     5221 gatatccttg atctgtggat ctaccacaca caaggctact tccctgatta gcagaactac 
     5281 acaccagggc caggggtcag atatccactg acctttggat ggtgctacaa gctagtacca 
     5341 gttgagccag ataaggtaga agaggccaat aaaggagaga acaccagctt gttacaccct 
     5401 gtgagcctgc atgggatgga tgacccggag agagaagtgt tagagtggag gtttgacagc 
     5461 cgcctagcat ttcatcacgt ggcccgagag ctgcatccgg agtacttcaa gaactgctga 
     5521 tatcgagctt gctacaaggg actttccgct ggggactttc cagggaggcg tggcctgggc 
     5581 gggactgggg agtggcgagc cctcagatcc tgcatataag cagctgcttt ttgcctgtac 
     5641 tgggtctctc tggttagacc agatctgagc ctgggagctc tctggctaac tagggaaccc 
     5701 actgcttaag cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt 
     5761 gtgtgactct ggtaactaga gatccctcag acccttttag tcagtgtgga aaatctctag 
     5821 cagtagtagt tcatgtcatc ttattattca gtatttataa cttgcaaaga aatgaatatc 
     5881 agagagtgag aggccttgac attgctagcg tttaccgtcg acctctagct agagcttggc 
     5941 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa 
     6001 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac 
     6061 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca 
     6121 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc 
     6181 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc 
     6241 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc 
     6301 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag 
     6361 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc 
     6421 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt 
     6481 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct 
     6541 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg 
     6601 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct 
     6661 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat 
     6721 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg 
     6781 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa 
     6841 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt 
     6901 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc 
     6961 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt 
     7021 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta 
     7081 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat 
     7141 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac 
     7201 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg 
     7261 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag 
     7321 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt 
     7381 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt 
     7441 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt 
     7501 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt 
     7561 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct 
     7621 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt 
     7681 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac 
     7741 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa 
     7801 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa 
     7861 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca 
     7921 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct 
     7981 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga 
     8041 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc 
     8101 tgacgtcgac ggatcgggag atcaacttgt ttattgcagc ttataatggt tacaaataaa 
     8161 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt 
     8221 tgtccaaact catcaatgta tcttatcatg tctggatcaa ctggataact caagctaacc 
     8281 aaaatcatcc caaacttccc accccatacc ctattaccac tgccaattac ctgtggtttc 
     8341 atttactcta aacctgtgat tcctctgaat tattttcatt ttaaagaaat tgtatttgtt 
     8401 aaatatgtac tacaaactta gtagttttta aagaaattgt atttgttaaa tatgtactac 
     8461 aaacttagta gt 
//

载体质粒图谱pdf版和相关资料下载

载体应用举例

分享到:

全部载体分类

相关载体

相关文章

相关问答

Copyright © 20012-2013 BIOFENG. 生物风 版权所有 Powered by Biofeng