The SDS gel was visualized with colloidal Coomassie blue stain (Invitrogen), imaged by Typhoon 9400 scanner followed by analysis with ImageQuant Software version TL 8.1 (GE Healthcare). Further processing of the proteins was then performed according to Thermo Scientific's TMT Mass Tagging Kits and...
The results were calculated by the 2−ΔΔCt method, and all the primer sequences required for qPCR in this study were shown in Table S1. Western blot analysis The samples were separated by SDS-PAGE and the protein was transferred to PVDF membrane (MilliporeSigma, Burlington, MA, USA)....
Protein Detection by Immunoblot—Standard procedures were used for yeast growth, cell harvesting, and cell breakage, as well as for preparation of protein-containing cell-free extracts, fractionation by SDS-polyacrylamide gel electrophoresis, and transfer to nitrocellulose membranes. Monoclonal anti-Myc (...
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Reactions were ended by the addition of SDS- PAGE loading buffer. Phosphorylation of MBP was analyzed by autoradiography after SDS-PAGE. The in-gel kinase activity assay was performed according to a procedure described previously [28] using 7 µg of total protein. Acknowledgments We would like...
Equal amounts of lysates (20–50 μg) were loaded on a Mini-Protean TGX (4%–20%) SDS-PAGE (Bio-Rad). The separated proteins were transferred onto polyvinylidene difluoride membranes (Amersham). For phospho-specific antibodies, the membranes were blocked for 1 hr with blocking buffer ...
以白菜型冬油菜"陇油6号"为材料,利用RT-PCR法克隆到全长MKK2基因cDNA,生物信息学分析表明油菜MKK2蛋白为亲水蛋白,包含有MAPKK蛋白所特有的磷酸化基序S/T-X3-5-S/T.将其与大肠杆菌表达载体pET30a连接,构建原核表达载体pET30a-MKK2,并转化大肠杆菌BL21,经IPTG诱导表达后,SDS-PAGE以及Western blotting检测结果表明...
Cell lysates were prepared and electrophoresed in SDS-PAGE and subsequently performed by Western blotting analysis. Data are expressed as mean ± SD (n = 3), aP < 0.05 vs the HepG2 group; cP < 0.05 vs pcDNA3.1-X transfected group. ...
取两组手术切除的肝癌组织及相应的癌旁正常组织(距肿瘤边缘>2 cm),RIPA-PMSF 充分匀浆,4 ℃ 10 000 r/min 离心 15 min,取上清。BCA 法 检测蛋白浓度合格后,加入上样缓冲液,95 ℃水浴 5 min;取蛋白 50 μg 行 SDS-PAGE 电泳(10%分离胶+5%浓缩胶),当蛋白跑至浓缩胶与分离胶交界时, 将电压由 80 ...
本发明涉及MEKK1‑MKK1/2‑MPK4级联对抗STOP1蛋白的磷酸化和稳定作用,具体地,本发明提供一种STOP1蛋白的磷酸化位点以及相应的磷酸化的STOP1蛋白,以及磷酸化的STOP1蛋白或其促进剂和/或MEKK1‑MKK1/2‑MPK4信号通路促进剂在提高植物的抗铝毒能力方面的用途。本发明首次发现,促进植物中STOP1蛋白的第386位...