| 出品公司: | Clontech |
|---|---|
| 载体名称: | pProLabel-C |
| 质粒类型: | 哺乳动物表达载体 |
| 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: | 4161 bp |
| 5' 测序引物及序列: | CMV Forward： CGCAAATGGGCGGTAGGCGTG |
| 3' 测序引物及序列: | -- |
| 载体标签: | ProLabel tag（N- ter） |
| 载体抗性: | 卡那霉素 |
| 筛选标记: | 新霉素（Neomycin） |
| 克隆菌株: | DH5α 等 |
| 宿主细胞（系）: | 常规细胞系（293、CV-1、CHO等） |
| 备注: | ProLabel tag 是一个分子量约6KD的标签，检测的灵敏度高。 |
| 产品目录号: | 630458 |
| 稳定性: | 稳表达 |
| 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
pProLabel-C载体描述
pProLabel-C is a mammalian expression vector designed to express a protein of interest fused to the the C-terminus of a 6 kDa ProLabel tag. The resulting fusion protein can be used in a variety of functional assays and quantitated with our ProLabel Detection Kit II (Cat. No. 631629). The Detection Kit provides all of the components needed to perform enzyme fragment complementation assays (1, 2). In these assays, two inactive enzyme fragments (the ProLabel tag, and a larger Enzyme Acceptor) are combined to form a complete, active enzyme that cleaves the Galacton Star chemiluminescent substrate. The resulting signal can be detected and quantitated with any standard luminometer.
The pProLabel-C vector contains a CMV promoter that drives strong, constitutive expression of the fusion protein, and an SV40 polyadenylation signal that directs processing of the 3’end of the mRNA transcript. The vector also contains a kanamycin/neomycin resistance cassette (Kanr/Neor) that allows G418 selection of stably transfected eukaryotic cells; a bacterial promoter upstream of this cassette allows kanamycin or neomycin selection of transformed bacterial cells. In addition, pProLabel-C contains an SV40 origin of replication for propagation in mammalian cells that express SV40 T-antigen, a pUC origin for propagation in E. coli and an f1 origin for the production of single-stranded DNA.
In order to create a fusion of your protein of interest and the ProLabel tag, your gene of interest must be in the same reading frame as the ProLabel tag sequence, and there can be no intervening stop codons. Prolabel vector constructs can be transfected into mammalian cells using standard transfection methods. ProLabel fusion protein expression levels can be measured quantitatively from mammalian cell lysates using the method described in the ProLabel Detection Kit II Protocol-at-a-Glance (PT3987-2). This highly sensitive and rapid procedure obviates the need for Western analysis. The assay is designed to be used with any ProLabel fusion in a variety of functional assays—from measuring target gene expression in RNAi knockdown studies to measuring protein interactions in coimmunoprecipitation studies.
质粒扩增方法
Suitable host strains: DH5α, Fusion Blue, and other general purpose strains. Single-stranded DNA production requires a host containing an F plasmid, such as JM101 or XL1-Blue.
Selectable marker: plasmid confers resistance to kanamycin (50 μg/ml) in E. coli hosts.
E. coli replication origin: pUC
Copy number: ~500
Plasmid incompatibility group: pMB1/ColE1
参考文献
1. Gorman, C. (1985) In DNA cloning: A practical approach, Vol. II. Ed. D.M. Glover. (IRL Press, Oxford, UK) pp. 143–190.
2. Eglen, R.M. and Singh, R. (2002) Comb. Chem. High Throughput Screen. 6: 381–387.
Note: The attached sequence file has been 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.
载体序列
LOCUS pProLabel-C 4161 bp DNA circular 1-FEB-2010
COMMENT Created by Clontech Laboratories Inc. http://www.clontech.com
COMMENT Cat. No. 630458,630461,630460,631628
FEATURES Location/Qualifiers
misc_feature 1..589
/label=CMV\IE\Promoter
misc_feature 613..780
/label=ProLabel\tag
misc_feature 782..825
/label=MCS
misc_feature 980..1030
/label=SV40\polyA\signals
misc_feature 1077..1532
/label=f1\origin
misc_feature 1873..2008
/label=SV40\origin
misc_feature 2057..2851
/label=Kan(R)/Neo(R)
misc_feature 3087..3105
/label=HSV\TK\polyA\signals
misc_feature 3436..4079
/label=pUC\origin
BASE COUNT 1002 a 1092 c 1074 g 993 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 ccggtcgcca ccatgagctc caattcactg gccgtcgttt tacaacgtcg tgactgggaa
661 aaccctggcg ttacccaact taatcgcctt gcagcacatc cccctttcgc cagctggcgt
721 aatagcgaag aggcccgcac cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa
781 caagcttcga attctgcagt cgacggtacc gcgggcccgg gatccaccgg atctagataa
841 ctgatcataa tcagccatac cacatttgta gaggttttac ttgctttaaa aaacctccca
901 cacctccccc tgaacctgaa acataaaatg aatgcaattg ttgttgttaa cttgtttatt
961 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
1021 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta acgcgtaaat
1081 tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt
1141 taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg
1201 gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt
1261 caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc
1321 aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg
1381 atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa
1441 aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc
1501 cgccgcgctt aatgcgccgc tacagggcgc gtcaggtggc acttttcggg gaaatgtgcg
1561 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
1621 ataaccctga taaatgcttc aataatattg aaaaaggaag agtcctgagg cggaaagaac
1681 cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga
1741 agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc
1801 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc
1861 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
1921 tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag
1981 aagtagtgag gaggcttttt tggaggccta ggcttttgca aagatcgatc aagagacagg
2041 atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg
2101 ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc
2161 cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg
2221 tgccctgaat gaactgcaag acgaggcagc gcggctatcg tggctggcca cgacgggcgt
2281 tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg
2341 cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat
2401 catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca
2461 ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca
2521 ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa
2581 ggcgagcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa
2641 tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc
2701 ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga
2761 atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc
2821 cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac
2881 caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg
2941 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
3001 atgctggagt tcttcgccca ccctaggggg aggctaactg aaacacggaa ggagacaata
3061 ccggaaggaa cccgcgctat gacggcaata aaaagacaga ataaaacgca cggtgttggg
3121 tcgtttgttc ataaacgcgg ggttcggtcc cagggctggc actctgtcga taccccaccg
3181 agaccccatt ggggccaata cgcccgcgtt tcttcctttt ccccacccca ccccccaagt
3241 tcgggtgaag gcccagggct cgcagccaac gtcggggcgg caggccctgc catagcctca
3301 ggttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
3361 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
3421 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
3481 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
3541 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
3601 actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
3661 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
3721 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
3781 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
3841 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
3901 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
3961 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
4021 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
4081 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
4141 aaccgtatta ccgccatgca t
//
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