| 出品公司: | Clontech |
|---|---|
| 载体名称: | pDsRED2-Mito |
| 质粒类型: | 哺乳细胞表达载体；荧光报告载体；亚细胞定位载体 |
| 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: | 4715bp |
| 5' 测序引物及序列: | -- |
| 3' 测序引物及序列: | -- |
| 载体标签: | 红色荧光蛋白DsRed2 |
| 载体抗性: | 卡纳霉素（Kanamycin） |
| 筛选标记: | 新霉素（Neomycin） |
| 备注: | pDsRED2-Mito载体是亚细胞定位载体，含有线粒体定位信号。 |
| 产品目录号: | 632421 |
| 稳定性: | 瞬表达 或 稳表达 |
| 组成型: | 组成型 |
| 病毒/非病毒: | 非病毒 |
买家导航
pDsRED2-Mito载体质粒基本信息
pDsRED2-Mito质粒图谱载体图谱和pDsRED2-Mito载体序列质粒序列多克隆位点信息
pDsRED2-Mito质粒载体简介
pDsRed2-Mito载体描述
fluorescent protein (DsRed2; 1, 2) and the mitochondrial targeting sequence from subunit VIII of human cytochrome c oxidase (Mito; 3, 4). The Mito sequence is fused to the 5'-end of DsRed2, a human codon-optimized DsRed variant that is engineered for faster maturation and lower nonspecific aggregation (1, 5). The Mito sequence targets the Mito-DsRed2 fusion protein to the host cell’s mitochondria.
To drive expression of Mito-DsRed2, this vector contains the immmediate early promoter of cytomegalovirus (PCMV IE ). SV40 polyadenylation signals downstream of the DsRed2 gene direct proper processing of the 3'-end of the Mito-DsRed2 mRNA. This vector also contains an SV40 origin for replication in any mammalian cell line that expresses the SV40 T-antigen, a pUC origin of replication for propagation in E. coli, and an f1 origin for single-stranded DNA production. A neomycin resistance cassette—consisting of the SV40 early promoter (PSV40e), the neomycin/kanamycin resistance gene of Tn5 (Neor/Kanr), and polyadenylation signals from the herpes simplex virus thymidine kinase (HSV TK poly A) gene—allow stably transfected eukaryotic cells to be selected using G418 (6). A bacterial promoter (P) upstream of this cassette drives expression of the gene encoding kanamycin resistance in E. coli.
pDsRed2-Mito is designed for fluorescent labeling of mitochondria. The vector can be introduced into mammalian cells using any standard transfection method. If required, stable transformants can be selected using G418 (6). The Mito-DsRed2 fusion (excitation/emission maxima: 558 nm/ 583 nm) can be detected by fluorescence microscopy and by flow cytometry. Filter sets optimized for detecting DsRed by microscopy are available from Chroma Technology Corporation and Omega Optical Inc. Please see their websites (www.chroma.com and www.omegafilters.com) and the Living Colors Vol. II User Manual, provided with this vector, for more information. To detect Mito- DsRed2-expressing cells by flow cytometry, use the instrument’s argon-ion laser to excite the fluorophore at 488 nm and the FL-2 channel to detect the fluorophore’s emission at 583 nm.
Propagation in E. coli
Suitable host strains: DH5α, HB101, and other general purpose strains. Single-stranded DNA production requires
a host containing an F plasmid such as JM109 or XL1-Blue.
Selectable marker: plasmid confers resistance to kanamycin (50 μg/ml) to E. coli hosts.
E. coli replication origin: pUC
Copy number: ~500
Plasmid incompatibility group: pMB1/Col E1
References
1. Living Colors DsRed2 (July 2001) Clontechniques XVI(3):2–3.
2. Matz, M. V., et al. (1999) Nature Biotech. 17:969–973.
3. Rizzuto, R., et al. (1995) Curr. Biol. 5:635–642.
4. Rizzuto, R., et al. (1989) J. Biol. Chem. 264:10595–10600.
5. Haas, J., et al. (1996) Curr. Biol. 6:315–324.
6. Gorman, C. (1985) In DNA Cloning: A Practical Approach, Vol. II, Ed. Glover, D. M. (IRL Press, Oxford, UK) pp. 143–190.
pDsRED2-Mito质粒序列载体序列
LOCUS pDsRed2-Mito 4715 bp DNA circular 8-JAN-2010
COMMENT Created by Clontech Laboratories Inc. http://www.biofeng.com/
COMMENT Cat. No. 632421
COMMENT VNTAUTHORNAME|biofeng.com|
FEATURES Location/Qualifiers
misc_feature 1..589
/label=CMV\IE\Promoter
misc_feature 597..683
/label=Mitochondrial\targeting\sequence
misc_feature 705..1382
/label=DsRed2
misc_feature 1534..1539
/label=SV40\early\polyA\signal
misc_feature 1563..1568
/label=SV40\early\polyA\signal
misc_feature 1631..2086
/label=f1\origin
misc_feature 2148..2176
/label=Bacterial\promoter\for\Kan(R)
misc_feature 2260..2489
/label=SV40\early\Promoter/Enhancer
misc_feature 2427..2562
/label=SV40\origin
misc_feature 2611..3405
/label=Kan(R)/Neo(R)
misc_feature 3641..3646
/label=HSV\TK\polyA\signal
misc_feature 3654..3659
/label=HSV\TK\polyA\signal
misc_feature 3990..4693
/label=pUC\origin
BASE COUNT 1105 a 1282 c 1260 g 1068 t
ORIGIN
1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcatgt
601 ccgtcctgac gccgctgctg ctgcggggct tgacaggctc ggcccggcgg ctcccagtgc
661 cgcgcgccaa gatccattcg ttgggggatc caccggtcgc caccatggcc tcctccgaga
721 acgtcatcac cgagttcatg cgcttcaagg tgcgcatgga gggcaccgtg aacggccacg
781 agttcgagat cgagggcgag ggcgagggcc gcccctacga gggccacaac accgtgaagc
841 tgaaggtgac caagggcggc cccctgccct tcgcctggga catcctgtcc ccccagttcc
901 agtacggctc caaggtgtac gtgaagcacc ccgccgacat ccccgactac aagaagctgt
961 ccttccccga gggcttcaag tgggagcgcg tgatgaactt cgaggacggc ggcgtggcga
1021 ccgtgaccca ggactcctcc ctgcaggacg gctgcttcat ctacaaggtg aagttcatcg
1081 gcgtgaactt cccctccgac ggccccgtga tgcagaagaa gaccatgggc tgggaggcct
1141 ccaccgagcg cctgtacccc cgcgacggcg tgctgaaggg cgagacccac aaggccctga
1201 agctgaagga cggcggccac tacctggtgg agttcaagtc catctacatg gccaagaagc
1261 ccgtgcagct gcccggctac tactacgtgg acgccaagct ggacatcacc tcccacaacg
1321 aggactacac catcgtggag cagtacgagc gcaccgaggg ccgccaccac ctgttcctgt
1381 agcggccgcg actctagatc ataatcagcc ataccacatt tgtagaggtt ttacttgctt
1441 taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca attgttgttg
1501 ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
1561 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
1621 cttaaggcgt aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
1681 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
1741 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
1801 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
1861 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
1921 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
1981 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
2041 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtcagg tggcactttt
2101 cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat
2161 ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtcct
2221 gaggcggaaa gaaccagctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct
2281 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa
2341 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
2401 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
2461 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct
2521 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaagatc
2581 gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt gcacgcaggt
2641 tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc
2701 tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag
2761 accgacctgt ccggtgccct gaatgaactg caagacgagg cagcgcggct atcgtggctg
2821 gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac
2881 tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc
2941 gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc
3001 tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc
3061 ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc agccgaactg
3121 ttcgccaggc tcaaggcgag catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat
3181 gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc
3241 cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa
3301 gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat
3361 tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt
3421 tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg
3481 ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc
3541 agcgcgggga tctcatgctg gagttcttcg cccaccctag ggggaggcta actgaaacac
3601 ggaaggagac aataccggaa ggaacccgcg ctatgacggc aataaaaaga cagaataaaa
3661 cgcacggtgt tgggtcgttt gttcataaac gcggggttcg gtcccagggc tggcactctg
3721 tcgatacccc accgagaccc cattggggcc aatacgcccg cgtttcttcc ttttccccac
3781 cccacccccc aagttcgggt gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc
3841 ctgccatagc ctcaggttac tcatatatac tttagattga tttaaaactt catttttaat
3901 ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg
3961 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
4021 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
4081 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
4141 cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact
4201 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg
4261 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc
4321 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
4381 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
4441 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
4501 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
4561 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
4621 ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc
4681 ctgattctgt ggataaccgt attaccgcca tgcat
//
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