- [10-03]如果PCR产物可以直接用来测序,但为什么还需要买PCR产物纯化试剂盒,还 ...
- [10-02]Egfr基因突变检测试剂盒及其应用
- [07-25]同源染色体和等位基因的联系和区别请问是不是有同源
- [08-03]点突变试剂盒/ QuikChange突变试剂盒系列
- [07-30]DNA测序仪的试剂器材
- [08-06]什么是白血病的靶向治疗?
- [08-04]TaKaRa 16S rDNA PCR扩增试剂盒说明书
- [07-21]线粒体提取试剂盒
- [07-27]建库试剂盒怎么选择?
Gen5 数据分析软件杭州生科仪器有限公司
蚂蚁淘
/发布时间:1630085406 /浏览量:326
SteveHahn
Lastmodified
Beforetryingthismethod,tryrapidproteinprepwhichinvolvesboilingyeastinSDSPAGEbuffer
1.Grow100mlyeasttoA600~1.0.Iftheyeastareovergrown,useproportionallylesscellsfortheprep.2.Collectcellsbycentrifugationandwashin20mlcoldextractionbuffer(containingDTTandproteaseinhibitors).Keepeverythingcoldfromthispointon.3.ResUSPendcellsin600microlitersextractionbufferandtransferto13x100mmglasstesttubesonice.Add400microlitersglassbeads.4.Inthecoldroom,vortexfor1minoniceandkeeponiceforatleast1minwhilevortexingothertubes.5.Repeatthevortexingstep(step4)foratotalof5times.6.Spincellsinprecooledclinicalcentrifugefor3minattopspeed.7.Usinga1mlpipetman,removethesupernatant,leavingmostoftheglassbeadsbehind(transferringsomeoftheyeastisOK).Transfertoamicrocentrifugetube.8.Inthecoldroom,spinattopspeedinthemicrofugefor30min.Removesupernatanttoanewtubebeingcarefulnottocarryoveranycelldebris.9.MeasureproteinconcentrationusingBioRadassayandfreezeinaliquots.Load20-40microgramsonproteingelforwesternanalysis.
ExtractionBuffer:200mMTrispH8.0150mMAmmoniumsulfate10%glycerol1mMEDTAAddfreshondayofextractpreparation:2mMDTTPMSFBenzamidineLeupeptinPepstatinChymostatinProteaseinhibitors0.1MPMSF(100x)16mg/mlEthanolStoreat-20degrees0.2MDTT(100x)32mg/mlH2OStorefrozenat-20degreesBenzamidine(100X)31mg/mlH2O.Storefrozenat-20degreesLeupeptin(500X)0.15mg/mlEthanol.Storeat-70degreesforlessthan6monthsPepstatin(200X)0.28mg/mlmethanolStoreat-20degrees.Chymostatin(2,500X)5mg/mlDMSOStorefrozenat-20degrees
================ 蚂蚁淘在线 ================
免责声明：本文仅代表作者个人观点，与本网无关。其创作性以及文中陈述文字和内容未经本站证实，对本文以及其中全部或者部分内容、文字的真实性、完整性、及时性本站不做任何保证或承诺，请读者仅作参考，并请自行核实相关内容
版权声明：未经蚂蚁淘在线授权不得转载、摘编或利用其他方式使用上述作品。已经经本网授权使用作品的，应该授权范围内使用，并注明“来源：蚂蚁淘在线”。违反上述声明者，本网将追究其相关法律责任。
