| 出品公司: | Clontech |
|---|---|
| 载体名称: | pCMV-tdTomato |
| 质粒类型: | 哺乳动物细胞表达载体；荧光报告载体 |
| 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: | CMV |
| 载体大小: | 5392 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: | 卡那霉素 |
| 筛选标记: | 新霉素（Neomycin） |
| 克隆菌株: | DH5α, HB101 |
| 宿主细胞（系）: | 常规细胞系（293、CV-1、CHO等） |
| 备注: | 哺乳动物载体pCMV-tdTomato组成型表达tdTomato蛋白。 |
| 产品目录号: | 632534 |
| 稳定性: | 稳表达 或 瞬表达 |
| 组成型/诱导型: | 组成型 |
| 病毒/非病毒: | 非病毒 |
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
pCMV-tdTomato载体描述
pCMV-tdTomato encodes tdTomato, a member of the family of fruit fluorescent proteins (1) derived from the Discosoma sp. red fluorescent protein, DsRed (2). Because the Tomato protein has a tendency to dimerize, the vector was designed with two copies of the Tomato coding region linked together to allow intramolecular dimerization. As a result, each tdTomato RNA transcript encodes a tandem dimer of the Tomato protein (excitation and emission maxima equal 554nm and 581nm, respectively). The tdTomato coding sequence is positioned just downstream of the constitutively active cytomegalovirus immediate early promoter (PCMV IE). As a result, mammalian cells transfected with this vector will constitutively express tdTomato. A Kozak consensus sequence is located immediately upstream of the tdTomato coding sequence to enhance translational efficiency in eukaryotic cells (3). SV40 polyadenylation signals downstream of the tdTomato coding sequence direct proper processing of the 3' end of the tdTomato mRNA. The vector backbone contains an SV40 origin for replication in mammalian cells expressing the SV40 large 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 (Neor) allows stably transfected eukaryotic cells to be selected using G418. This cassette consists of the SV40 early promoter, the neomycin/kanamycin resistance gene of Tn5, and polyadenylation signals from the Herpes simplex virus thymidine kinase (HSV TK) gene. A bacterial promoter upstream of the cassette expresses kanamycin resistance in E. coli.
pCMV-tdTomato is designed for use as a marker for cotransfection or cell tracking. The red fluorescence of tdTomato can be detected by fluorescence microscopy, allowing direct visual imaging. In addition, flow cytometry can be used to enrich transfected cell populations. pCMV-tdTomato can be transfected into mammalian cells using any standard transfection method. If required, stable transfectants can be selected using G418 (4).
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
Excitation and emission maxima of tdTomato
Excitation maximum = 554 nm
Emission maximum = 581 nm
References
1. Shaner, N. C., et al. (2004) Nature Biotech. 22(12):1567-1572.
2. Bevis, B. J. & Glick, B. S. (2002) Nature Biotech. 20:83–87.
3. Kozak, M. (1987) Nucleic Acids Res. 15:8125–8148.
4. Gorman, C. (1985) In DNA cloning: A Practical Approach, Vol. II. Ed. D. M. Glover. (IRL Press, Oxford, U.K.), pp. 143–190.
载体序列
LOCUS pCMV-td-Tomato 5392 bp DNA circular 28-JUL-2008
COMMENT Created by Clontech Laboratories Inc. http://www.clontech.com
COMMENT Cat. No. 632534
FEATURES Location/Qualifiers
misc_feature 3288..4082
/label=KanR/NeoR
CDS 628..2055
/label=td-Tomato
promoter 1..589
/label=pCMV\IE
BASE COUNT 1272 a 1505 c 1459 g 1156 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 gctagcgcta
601 ccggactcag atccaccggt cgccaccatg gtgagcaagg gcgaggaggt catcaaagag
661 ttcatgcgct tcaaggtgcg catggagggc tccatgaacg gccacgagtt cgagatcgag
721 ggcgagggcg agggccgccc ctacgagggc acccagaccg ccaagctgaa ggtgaccaag
781 ggcggccccc tgcccttcgc ctgggacatc ctgtcccccc agttcatgta cggctccaag
841 gcgtacgtga agcaccccgc cgacatcccc gattacaaga agctgtcctt ccccgagggc
901 ttcaagtggg agcgcgtgat gaacttcgag gacggcggtc tggtgaccgt gacccaggac
961 tcctccctgc aggacggcac gctgatctac aaggtgaaga tgcgcggcac caacttcccc
1021 cccgacggcc ccgtaatgca gaagaagacc atgggctggg aggcctccac cgagcgcctg
1081 tacccccgcg acggcgtgct gaagggcgag atccaccagg ccctgaagct gaaggacggc
1141 ggccactacc tggtggagtt caagaccatc tacatggcca agaagcccgt gcaactgccc
1201 ggctactact acgtggacac caagctggac atcacctccc acaacgagga ctacaccatc
1261 gtggaacagt acgagcgctc cgagggccgc caccacctgt tcctggggca tggcaccggc
1321 agcaccggca gcggcagctc cggcaccgcc tcctccgagg acaacaacat ggccgtcatc
1381 aaagagttca tgcgcttcaa ggtgcgcatg gagggctcca tgaacggcca cgagttcgag
1441 atcgagggcg agggcgaggg ccgcccctac gagggcaccc agaccgccaa gctgaaggtg
1501 accaagggcg gccccctgcc cttcgcctgg gacatcctgt ccccccagtt catgtacggc
1561 tccaaggcgt acgtgaagca ccccgccgac atccccgatt acaagaagct gtccttcccc
1621 gagggcttca agtgggagcg cgtgatgaac ttcgaggacg gcggtctggt gaccgtgacc
1681 caggactcct ccctgcagga cggcacgctg atctacaagg tgaagatgcg cggcaccaac
1741 ttcccccccg acggccccgt aatgcagaag aagaccatgg gctgggaggc ctccaccgag
1801 cgcctgtacc cccgcgacgg cgtgctgaag ggcgagatcc accaggccct gaagctgaag
1861 gacggcggcc actacctggt ggagttcaag accatctaca tggccaagaa gcccgtgcaa
1921 ctgcccggct actactacgt ggacaccaag ctggacatca cctcccacaa cgaggactac
1981 accatcgtgg aacagtacga gcgctccgag ggccgccacc acctgttcct gtacggcatg
2041 gacgagctgt acaagtaggc ggccgcgact ctagatcata atcagccata ccacatttgt
2101 agaggtttta cttgctttaa aaaacctccc acacctcccc ctgaacctga aacataaaat
2161 gaatgcaatt gttgttgtta acttgtttat tgcagcttat aatggttaca aataaagcaa
2221 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
2281 caaactcatc aatgtatctt aaggcgtaaa ttgtaagcgt taatattttg ttaaaattcg
2341 cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc
2401 cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga
2461 gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg
2521 atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg tgccgtaaag
2581 cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga
2641 acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg
2701 tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg
2761 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
2821 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
2881 gaaaaaggaa gagtcctgag gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg
2941 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
3001 agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
3061 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc
3121 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
3181 cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
3241 aggcttttgc aaagatcgat caagagacag gatgaggatc gtttcgcatg attgaacaag
3301 atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg
3361 cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc
3421 cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcaa gacgaggcag
3481 cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca
3541 ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat
3601 ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata
3661 cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac
3721 gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc
3781 tcgcgccagc cgaactgttc gccaggctca aggcgagcat gcccgacggc gaggatctcg
3841 tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg
3901 gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta
3961 cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg
4021 gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct
4081 gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga
4141 tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc
4201 cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc accctagggg
4261 gaggctaact gaaacacgga aggagacaat accggaagga acccgcgcta tgacggcaat
4321 aaaaagacag aataaaacgc acggtgttgg gtcgtttgtt cataaacgcg gggttcggtc
4381 ccagggctgg cactctgtcg ataccccacc gagaccccat tggggccaat acgcccgcgt
4441 ttcttccttt tccccacccc accccccaag ttcgggtgaa ggcccagggc tcgcagccaa
4501 cgtcggggcg gcaggccctg ccatagcctc aggttactca tatatacttt agattgattt
4561 aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac
4621 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa
4681 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
4741 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
4801 aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg
4861 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
4921 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
4981 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
5041 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
5101 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
5161 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
5221 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
5281 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt
5341 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgccatgc at
//
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