| 出品公司: | Clontech |
|---|---|
| 载体名称: | pLVX-IRES-tdTomato |
| 质粒类型: | 慢病毒载体；哺乳动物细胞表达载体；双顺反子载体；荧光报告载体 |
| 高拷贝/低拷贝: | 高拷贝 |
| 启动子: | CMV |
| 克隆方法: | 多克隆位点，限制性内切酶 |
| 载体大小: | 8892 bp |
| 5' 测序引物及序列: | CMV-F: CGCAAATGGGCGGTAGGCGTG (Invitrogen) |
| 3' 测序引物及序列: | -- |
| 载体标签: | 无标签 |
| 载体抗性: | 氨苄青霉素 |
| 筛选标记: | 嘌呤霉素(Puromycin) |
| 克隆菌株: | Stbl3 |
| 备注: |
pLVX-IRES-tdTomato载体含IRES（内部核糖体结合位点）元件； 目的基因和tdTomato荧光蛋白基因以双顺反子形式表达； CMV启动子驱动目的基因的过表达； tdTomato是第三代荧光蛋白，亮度高，适用于流式细胞仪。 |
| 产品目录号: | 631238 |
| 稳定性: | 稳表达 |
| 组成型: | 组成型 |
| 病毒/非病毒: | 慢病毒 |
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
Description
pLVX-IRES-tdTomato is an HIV-1-based, lentiviral expression vector that allows the simultaneous expression of your protein of interest and tdTomato in virtually any mammalian cell type, including primary cells. tdTomato is a member of the family of fruit fluorescent proteins derived from the Discosoma sp. red fluorescent protein, DsRed (1). The vector expresses the two proteins from a bicistronic mRNA transcript, allowing tdTomato to be used as an indicator of transduction efficiency and a marker for selection by flow cytometry.
Expression of the bicistronic transcript is driven by the constitutively active human cytomegalovirus immediate early promoter (PCMV IE) located just upstream of the MCS. An encephalomyocarditis virus (EMCV) internal ribosome entry site (IRES), positioned between the MCS and tdTomato, facilitates cap-independent translation of tdTomato from an internal start site at the IRES/tdTomato junction (1).
pLVX-IRES-tdTomato 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 RNA (2), leading to increased viral titers from packaging 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-IRES-tdTomato 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). The vector also contains a pUC origin of replication and an E. coli ampicillin resistance gene (Ampr) for propagation and selection in bacteria.
Use
pLVX-IRES-tdTomato is designed to constitutively coexpress your protein of interest and tdTomato from PCMV IE when transduced into mammalian cells. Before it can be transduced into target cells, the vector must be packaged into viral particles in HEK293T cells, using our Lenti-X™ HT Packaging System (Cat. Nos. 632160 and 632161). 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 nondividing and primary cells (5).
The presence of tdTomato allows transductants to be visualized by fluorescence microscopy and sorted by flow cytometry with standard FITC filter sets (tdTomato has an excitation maximum of 554 nm and an emission maximum of 581 nm).
载体序列
LOCUS pLVX-IRES-tdTomato 8892 bp DNA circular 2-JUN-2009
COMMENT Created by Clontech Laboratories Inc.
COMMENT Cat. No. 631238
COMMENT This file is created by Vector NTI
http://www.biofeng.com/
COMMENT VNTAUTHORNAME|biofeng.com|
FEATURES Location/Qualifiers
LTR 1..635
/label=5'\LTR
misc_feature 685..822
/label=Packaging\Signal
misc_binding 1303..1536
/label=RRE
promoter 2185..2787
/label=CMV\IE\Promoter
misc_feature 2842..3416
/label=IRES
CDS 3417..4844
/label=tdTomato
misc_feature 4861..5452
/label=WPRE
LTR 5655..6291
/label=3'\LTR
rep_origin complement(6760..7433)
/label=pUC\ori
CDS complement(7578..8574)
/label=Amp(R)
misc_feature 636..653
/label=PBS
misc_feature 2028..2151
/label=cPPT/CTS\(DNA\flap)
misc_feature 2803..2840
/label=MCS
BASE COUNT 2285 a 2247 c 2310 g 2050 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 tctatttccg gtgaattcct cgagactagt
2821 tctagagcgg ccgcggatcc cgcccctctc cctccccccc ccctaacgtt actggccgaa
2881 gccgcttgga ataaggccgg tgtgcgtttg tctatatgtt attttccacc atattgccgt
2941 cttttggcaa tgtgagggcc cggaaacctg gccctgtctt cttgacgagc attcctaggg
3001 gtctttcccc tctcgccaaa ggaatgcaag gtctgttgaa tgtcgtgaag gaagcagttc
3061 ctctggaagc ttcttgaaga caaacaacgt ctgtagcgac cctttgcagg cagcggaacc
3121 ccccacctgg cgacaggtgc ctctgcggcc aaaagccacg tgtataagat acacctgcaa
3181 aggcggcaca accccagtgc cacgttgtga gttggatagt tgtggaaaga gtcaaatggc
3241 tcacctcaag cgtattcaac aaggggctga aggatgccca gaaggtaccc cattgtatgg
3301 gatctgatct ggggcctcgg tgcacatgct ttacatgtgt ttagtcgagg ttaaaaaacg
3361 tctaggcccc ccgaaccacg gggacgtggt tttcctttga aaaacacgat gataatatgg
3421 tgagcaaggg cgaggaggtc atcaaagagt tcatgcgctt caaggtgcgc atggagggct
3481 ccatgaacgg ccacgagttc gagatcgagg gcgagggcga gggccgcccc tacgagggca
3541 cccagaccgc caagctgaag gtgaccaagg gcggccccct gcccttcgcc tgggacatcc
3601 tgtcccccca gttcatgtac ggctccaagg cgtacgtgaa gcaccccgcc gacatccccg
3661 attacaagaa gctgtccttc cccgagggct tcaagtggga gcgcgtgatg aacttcgagg
3721 acggcggtct ggtgaccgtg acccaggact cctccctgca ggacggcacg ctgatctaca
3781 aggtgaagat gcgcggcacc aacttccccc ccgacggccc cgtaatgcag aagaagacca
3841 tgggctggga ggcctccacc gagcgcctgt acccccgcga cggcgtgctg aagggcgaga
3901 tccaccaggc cctgaagctg aaggacggcg gccactacct ggtggagttc aagaccatct
3961 acatggccaa gaagcccgtg caactgcccg gctactacta cgtggacacc aagctggaca
4021 tcacctccca caacgaggac tacaccatcg tggaacagta cgagcgctcc gagggccgcc
4081 accacctgtt cctggggcat ggcaccggca gcaccggcag cggcagctcc ggcaccgcct
4141 cctccgagga caacaacatg gccgtcatca aagagttcat gcgcttcaag gtgcgcatgg
4201 agggctccat gaacggccac gagttcgaga tcgagggcga gggcgagggc cgcccctacg
4261 agggcaccca gaccgccaag ctgaaggtga ccaagggcgg ccccctgccc ttcgcctggg
4321 acatcctgtc cccccagttc atgtacggct ccaaggcgta cgtgaagcac cccgccgaca
4381 tccccgatta caagaagctg tccttccccg agggcttcaa gtgggagcgc gtgatgaact
4441 tcgaggacgg cggtctggtg accgtgaccc aggactcctc cctgcaggac ggcacgctga
4501 tctacaaggt gaagatgcgc ggcaccaact tcccccccga cggccccgta atgcagaaga
4561 agaccatggg ctgggaggcc tccaccgagc gcctgtaccc ccgcgacggc gtgctgaagg
4621 gcgagatcca ccaggccctg aagctgaagg acggcggcca ctacctggtg gagttcaaga
4681 ccatctacat ggccaagaag cccgtgcaac tgcccggcta ctactacgtg gacaccaagc
4741 tggacatcac ctcccacaac gaggactaca ccatcgtgga acagtacgag cgctccgagg
4801 gccgccacca cctgttcctg tacggcatgg acgagctgta caagtgaacg cgtctggaac
4861 aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct
4921 ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt
4981 atggctttca ttttctcctc cttgtataaa tcctggttgc tgtctcttta tgaggagttg
5041 tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc aacccccact
5101 ggttggggca ttgccaccac ctgtcagctc ctttccggga ctttcgcttt ccccctccct
5161 attgccacgg cggaactcat cgccgcctgc cttgcccgct gctggacagg ggctcggctg
5221 ttgggcactg acaattccgt ggtgttgtcg gggaagctga cgtcctttcc atggctgctc
5281 gcctgtgttg ccacctggat tctgcgcggg acgtccttct gctacgtccc ttcggccctc
5341 aatccagcgg accttccttc ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt
5401 cgccttcgcc ctcagacgag tcggatctcc ctttgggccg cctccccgcc tggaattaat
5461 tctgcagtcg agacctagaa aaacatggag caatcacaag tagcaataca gcagctacca
5521 atgctgattg tgcctggcta gaagcacaag aggaggagga ggtgggtttt ccagtcacac
5581 ctcaggtacc tttaagacca atgacttaca aggcagctgt agatcttagc cactttttaa
5641 aagaaaagag gggactggaa gggctaattc actcccaacg aagacaagat atccttgatc
5701 tgtggatcta ccacacacaa ggctacttcc ctgattagca gaactacaca ccagggccag
5761 gggtcagata tccactgacc tttggatggt gctacaagct agtaccagtt gagccagata
5821 aggtagaaga ggccaataaa ggagagaaca ccagcttgtt acaccctgtg agcctgcatg
5881 ggatggatga cccggagaga gaagtgttag agtggaggtt tgacagccgc ctagcatttc
5941 atcacgtggc ccgagagctg catccggagt acttcaagaa ctgctgatat cgagcttgct
6001 acaagggact ttccgctggg gactttccag ggaggcgtgg cctgggcggg actggggagt
6061 ggcgagccct cagatcctgc atataagcag ctgctttttg cctgtactgg gtctctctgg
6121 ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact gcttaagcct
6181 caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt
6241 aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag tagtagttca
6301 tgtcatctta ttattcagta tttataactt gcaaagaaat gaatatcaga gagtgagagg
6361 ccttgacatt gctagcgttt accgtcgacc tctagctaga gcttggcgta atcatggtca
6421 tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga
6481 agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg
6541 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc
6601 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac
6661 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
6721 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
6781 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
6841 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
6901 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
6961 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
7021 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
7081 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
7141 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
7201 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga
7261 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
7321 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
7381 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
7441 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
7501 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
7561 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
7621 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
7681 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
7741 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
7801 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
7861 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
7921 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
7981 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
8041 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
8101 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
8161 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
8221 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
8281 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
8341 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
8401 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
8461 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
8521 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcgacgga
8581 tcgggagatc aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
8641 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
8701 caatgtatct tatcatgtct ggatcaactg gataactcaa gctaaccaaa atcatcccaa
8761 acttcccacc ccatacccta ttaccactgc caattacctg tggtttcatt tactctaaac
8821 ctgtgattcc tctgaattat tttcatttta aagaaattgt atttgttaaa tatgtactac
8881 aaacttagta gt
//
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