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Ask a QuestionSupport@biorbyt.comRat CD133 Human Cells Transfected Lysate (positive control) (denatured)
Catalog Number: orb174679
Product Name | Rat CD133 Human Cells Transfected Lysate (positive control) (denatured) |
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Catalog Number | orb174679 |
Hazard Information | Non-Toxic |
Target | Rat CD133 |
Form/Appearance | 50 mM Tris-HCl pH 6.8, 12.5% glycerol, 1% sodium dodecylsulfate, 0.01% bromophenol blue |
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Storage | Store at -80°C. Aliquot to avoid repeated freezing and thawing |
Note | For research use only. |
Biological Origin | Human Cells |
Human Cells transfected lysate in which Rat CD133/PROM1/Prominin 1 has been over-expressed. The whole cell lysate is provided in 1X Sample Buffer.
Application Notes | Cell lysate was prepared by homogenization in ice-cold modified RIPA Lysis Buffer with cocktail of protease inhibitors (Sigma). Cell debris was removed by centrifugation. Protein concentration was determined with Bradford assay (Bio-Rad protein assay, Microplate Standard assay). The cell lysate was boiled for 5 minutes in 1 x SDS sample buffer (50 mM Tris-HCl pH 6.8, 12.5% glycerol, 1% sodium dodecylsulfate, 0.01% bromophenol blue) containing 5% b-mercaptoethanol |
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Material Safety Data Sheet
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先提取RNA,反转录成cDNA,然后根据目的基因设计PCR引物,通过半定量RT-PCR确定目的基因表达.
正因为检测的是mRNA,所以要先反转录成cDNA才能PCR.
① 在构建载体时,目的基因直接整合到细胞染色体组上,最好不要通过先瞬转在筛选稳定细胞株的这种方法,因为转染效率没有保证
② 高表达载体的构建,哺乳动物表达量一直是它自身的缺点,最好根据高表达载体定向的驯化细胞,提高蛋白表达量
③ 细胞的选择,筛选稳定细胞株我们常用的细胞是CHO,中国仓鼠卵巢细胞,由于CHO具有诸多的优点因此适合用于筛选稳定细胞株,而HEK293细胞则常用于瞬时转染
④ 后期的筛选,双抗预防污染,筛选细胞的时候抗生素浓度一定要做预实验,而且转染的时候不能有抗生素,关于细胞转染 稳定细胞系构建的相关理论
最近因工作需要,欲购进一些生产用的细胞株和瞬转、稳转质粒,大家有什么推荐的品系和公司或网站,谢谢!!!
主要需求:
1.生产或生物制药用的,稳定细胞株如CHO-K1、GS和HEK293/HEK293T,以及与之对应的瞬转、稳转(重组整合)质粒(需详细图谱)。
2.原始或改造细胞株,以及对应质粒,改造细胞株请详述细节,原因,优缺点,安全性等。
3.希望大家积极推荐,实验用、生产用均可,原核、真核都行,网站或品系名称也行,只要大家觉得常用、稳定、安全、高产就行。
4.请有意向的同仁回帖或将详细资料发至我163邮箱:wangqiang23mars@163.com,万分感谢。
然后你得确定,这个细胞株对于你得研究来说不可或缺,具有明确的代表性。
上面两个都没有问题的话,祝你顺利。
如果经费充足的话可以找公司包装病毒,如武汉的普健可以提供各种载体的构建及细胞株构建的技术服务。
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