pLVX-MetLuc

pLVX-MetLuc

载体基本信息

出品公司:	Ybscience
载体名称:	pLVX-MetLuc
质粒类型:	慢病毒载体；无启动子载体；荧光素酶报告载体
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:	无
载体大小:	8088 bp
5' 测序引物及序列:	--
3' 测序引物及序列:	--
载体标签:	--
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:	慢病毒载体pLVX-MetLuc没有启动子元件； MetLuc是已知最小分子量的荧光素酶； 荧光素酶报告基因可以用来定性定量分析启动子及其它顺式转录元件的活 性。
稳定性:	稳表达
组成型/诱导型:	--
病毒/非病毒:	慢病毒

 

载体质粒图谱和多克隆位点信息

载体简介

 

pLVX-MetLuc载体描述

pLVX-MetLuc is a promoterless, HIV-1-based, lentiviral vector that allows the functional analysis of different promoters and promoter/enhancer combinations inserted into its multiple cloning site (MCS). The vector encodes Metridia luciferase (MetLuc), a secreted reporter protein that can be easily detected in the medium surrounding the cells.
Metridia luciferase is a naturally secreted luciferase from the marine copepod Metridia longa. The human codon-optimized Metridia luciferase gene encodes a 219 amino acid (24 kDa) polypeptide that includes a 17 amino acid N-terminal signal peptide necessary for secretion (1). When a functional promoter is cloned into the MCS, located immediately upstream of the reporter, Metridia luciferase is expressed and secreted into the surrounding medium, where it is easily detected by the addition of a chemiluminescent substrate.
Lentiviral particles derived from pLVX-MetLuc allow you to monitor the activity of your promoter of interest in virtually any cell type, even primary cells. pLVX-MetLuc 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 (2), 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 (3). Finally, pLVX-MetLuc 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 (4).In addition to lentiviral elements, pLVX-MetLuc contains a puromycin resistance gene (Puror) under the control of the murine phosphoglycerate kinase promoter (PPGK) for  selection of stable transductants. The vector also contains a pUC origin of replication and an E. coli ampicillin resistance gene (Ampr) for propagation and selection in bacteria.

The pLVX-MetLuc vector, available as part of the Lenti-X Ready-To-Glow Secreted Luciferase Reporter System (Cat. No. 631746), can be used to monitor promoter activity in live cells as well as in vivo. A promoter must be cloned into the MCS, located upstream of the Metridia luciferase coding sequence. Without the addition of a functional promoter, the vector will not express MetLuc. Lentiviral particles derived from the vector allow you to monitor your promoter of interest in virtually any cell type, even primary cells. If required, stable transfectants can be selected using puromycin.

Propagation in E. coli
 Recommended host strains: DH5α and other general purpose strains.
 Selectable marker: plasmid confers resistance to ampicillin (100 μg/ml) in E. coli hosts.
 E. coli replication origin: pUC

References
1. Markova, S.V. et al. (2004) J. Bio. Chem. 279(5):3212-3217.
2. Zufferey, R. et al. (1999) J. Virol. 73(4):2886–2892.
3. Cochrane, A. W. et al. (1990) Proc. Natl. Acad. Sci. USA 87(3):1198–1202.
4. Zennou, V. et al. (2000) Cell 101(2):173–185.

 

载体序列

LOCUS       pLVX-MetLuc_Reporter   8088 bp    DNA   circular             6-NOV-2009
COMMENT     Created by Clontech Laboratories Inc. http://www.clontech.com
COMMENT     Cat. No. 631747,631746
FEATURES             Location/Qualifiers
     LTR             1..635
                     /label=5'\LTR
     misc_feature    685..822
                     /label=packaging\signal
     misc_feature    4056..4647
                     /label=WPRE
     rep_origin      complement(5956..6629)
                     /label=pUC\origin
     CDS             complement(6774..7770)
                     /label=Amp(R)
     misc_binding    1303..1536
                     /label=RRE
     misc_binding    636..653
                     /label=PBS
     misc_feature    2028..2151
                     /label=cPPT/CTS
     LTR             4850..5486
                     /label=3'\LTR
     misc_feature    2914..3422
                     /label=PGK\promoter
     misc_feature    3443..4042
                     /label=Puro(R)
     misc_feature    2195..2234
                     /label=MCS
     CDS             2247..2903
                     /label=Metridia\luciferase
BASE COUNT     2019 a      2085 c      2157 g      1827 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 cgtcgagctc aagcttcgaa ttctgcagtc gacggtaccg 
     2221 cgggcccggg atcccgcgac tctagaatgg acatcaaggt ggtgttcacc ctggtgttca 
     2281 gcgccctggt gcaggccaag agcaccgagt tcgaccccaa catcgacatc gtgggcctgg 
     2341 aaggcaagtt cggcatcacc aacctggaaa ccgacctgtt caccatctgg gagaccatgg 
     2401 aagtgatgat caaggccgac atcgccgaca ccgaccgggc cagcaacttc gtggccaccg 
     2461 agaccgacgc caaccggggc aagatgcccg gcaagaagct gcccctggcc gtcatcatgg 
     2521 aaatggaagc caacgccttc aaggccggct gcacccgggg ctgcctgatc tgcctgagca 
     2581 agatcaagtg caccgccaag atgaaggtgt acatccccgg caggtgccac gactacggcg 
     2641 gcgacaagaa aaccggccag gccggcatcg tgggcgccat cgtggacatc cccgagatca 
     2701 gcggcttcaa agaaatggcc cccatggaac agttcatcgc ccaggtggac agatgcgcca 
     2761 gctgcaccac cggctgcctg aagggcctgg ccaacgtgaa gtgcagcgag ctgctgaaga 
     2821 agtggctgcc cgaccgctgc gccagcttcg ccgacaagat ccagaaagag gtgcacaaca 
     2881 tcaagggcat ggccggcgac aggtagtcta gataattcta ccgggtaggg gaggcgcttt 
     2941 tcccaaggca gtctggagca tgcgctttag cagccccgct gggcacttgg cgctacacaa 
     3001 gtggcctctg gcctcgcaca cattccacat ccaccggtag gcgccaaccg gctccgttct 
     3061 ttggtggccc cttcgcgcca ccttctactc ctcccctagt caggaagttc ccccccgccc 
     3121 cgcagctcgc gtcgtgcagg acgtgacaaa tggaagtagc acgtctcact agtctcgtgc 
     3181 agatggacag caccgctgag caatggaagc gggtaggcct ttggggcagc ggccaatagc 
     3241 agctttgctc cttcgctttc tgggctcaga ggctgggaag gggtgggtcc gggggcgggc 
     3301 tcaggggcgg gctcaggggc ggggcgggcg cccgaaggtc ctccggaggc ccggcattct 
     3361 gcacgcttca aaagcgcacg tctgccgcgc tgttctcctc ttcctcatct ccgggccttt 
     3421 cgacctgcag cccaagctta ccatgaccga gtacaagccc acggtgcgcc tcgccacccg 
     3481 cgacgacgtc cccagggccg tacgcaccct cgccgccgcg ttcgccgact accccgccac 
     3541 gcgccacacc gtcgatccgg accgccacat cgagcgggtc accgagctgc aagaactctt 
     3601 cctcacgcgc gtcgggctcg acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt 
     3661 ggcggtctgg accacgccgg agagcgtcga agcgggggcg gtgttcgccg agatcggccc 
     3721 gcgcatggcc gagttgagcg gttcccggct ggccgcgcag caacagatgg aaggcctcct 
     3781 ggcgccgcac cggcccaagg agcccgcgtg gttcctggcc accgtcggcg tctcgcccga 
     3841 ccaccagggc aagggtctgg gcagcgccgt cgtgctcccc ggagtggagg cggccgagcg 
     3901 cgccggggtg cccgccttcc tggagacctc cgcgccccgc aacctcccct tctacgagcg 
     3961 gctcggcttc accgtcaccg ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat 
     4021 gacccgcaag cccggtgcct gaccgcgtct ggaacaatca acctctggat tacaaaattt 
     4081 gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg 
     4141 ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt 
     4201 ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg 
     4261 tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc 
     4321 agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg 
     4381 cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt 
     4441 tgtcggggaa gctgacgtcc tttccatggc tgctcgcctg tgttgccacc tggattctgc 
     4501 gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg 
     4561 gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag acgagtcgga 
     4621 tctccctttg ggccgcctcc ccgcctggaa ttaattctgc agtcgagacc tagaaaaaca 
     4681 tggagcaatc acaagtagca atacagcagc taccaatgct gattgtgcct ggctagaagc 
     4741 acaagaggag gaggaggtgg gttttccagt cacacctcag gtacctttaa gaccaatgac 
     4801 ttacaaggca gctgtagatc ttagccactt tttaaaagaa aagaggggac tggaagggct 
     4861 aattcactcc caacgaagac aagatatcct tgatctgtgg atctaccaca cacaaggcta 
     4921 cttccctgat tagcagaact acacaccagg gccaggggtc agatatccac tgacctttgg 
     4981 atggtgctac aagctagtac cagttgagcc agataaggta gaagaggcca ataaaggaga 
     5041 gaacaccagc ttgttacacc ctgtgagcct gcatgggatg gatgacccgg agagagaagt 
     5101 gttagagtgg aggtttgaca gccgcctagc atttcatcac gtggcccgag agctgcatcc 
     5161 ggagtacttc aagaactgct gatatcgagc ttgctacaag ggactttccg ctggggactt 
     5221 tccagggagg cgtggcctgg gcgggactgg ggagtggcga gccctcagat cctgcatata 
     5281 agcagctgct ttttgcctgt actgggtctc tctggttaga ccagatctga gcctgggagc 
     5341 tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct tgagtgcttc 
     5401 aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc agaccctttt 
     5461 agtcagtgtg gaaaatctct agcagtagta gttcatgtca tcttattatt cagtatttat 
     5521 aacttgcaaa gaaatgaata tcagagagtg agaggccttg acattgctag cgttttaccg 
     5581 tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt 
     5641 tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt 
     5701 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg 
     5761 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg 
     5821 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg 
     5881 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat 
     5941 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc 
     6001 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc 
     6061 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga 
     6121 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt 
     6181 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg 
     6241 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc 
     6301 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg 
     6361 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc 
     6421 ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg 
     6481 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc 
     6541 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct 
     6601 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt 
     6661 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa 
     6721 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa 
     6781 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc 
     6841 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct 
     6901 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca 
     6961 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt 
     7021 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt 
     7081 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc 
     7141 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc 
     7201 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt 
     7261 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact 
     7321 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc 
     7381 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt 
     7441 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg 
     7501 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct 
     7561 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa 
     7621 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt 
     7681 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc 
     7741 acatttcccc gaaaagtgcc acctgacgtc gacggatcgg gagatcaact tgtttattgc 
     7801 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt 
     7861 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctggat 
     7921 caactggata actcaagcta accaaaatca tcccaaactt cccaccccat accctattac 
     7981 cactgccaat tacctgtggt ttcatttact ctaaacctgt gattcctctg aattattttc 
     8041 attttaaaga aattgtattt gttaaatatg tactacaaac ttagtagt