梅花鹿鹿茸不同部位蛋白双向电泳谱图比较

目的:建立一套适合梅花鹿鹿茸蛋白质组学研究的双向电泳方法,并以此为基础比较不同部位梅花鹿鹿茸的蛋白图谱差异。方法:使用含7 mol/L尿素、4%CHAPS、2 mol/L硫脲、65 mmol/L DTT、1 mmol/L PMSF和0.2%Bio-Lyte的溶液为蛋白裂解液提取鹿茸不同部位的总蛋白,采用固相pH 3～10线性梯度,7 cm胶条进行第一向聚焦,并对等电聚焦参数进行优化,12%SDS聚丙烯凝胶进行第二向电泳,得到的凝胶经考马斯亮蓝染色后用PDQuest软件进行图谱比较分析。结果:3个部位总蛋白含量及蛋白点数从上到下依次降低,将3个图谱经软件分析获得18个差异蛋白点,同时初步得到每个蛋白点的等电点、分子量以及灰度值。结论:优化了适合梅花鹿鹿茸蛋白的双向电泳方法;不同部位鹿茸蛋白双向电泳图谱存在明显差异。

人成熟卵泡液蛋白质谱图的初步研究

目的:建立人成熟卵泡液的蛋白质谱图,筛选鉴定卵泡发育/卵母细胞成熟的相关蛋白质。方法:选取48例因男性因素行ICSI-ET治疗的女性为研究对象,收集取卵时获得的人成熟卵泡液,采用双向凝胶电泳-质谱技术分离鉴定人成熟卵泡液蛋白质谱图。结果:获得了较高分辨率的人成熟卵泡液蛋白质谱图,共检测到180多个蛋白质点,筛选鉴定了5个蛋白质点,分别是载脂蛋白E、PRO2044、Hypothetical protein、免疫球蛋白G重链以及转铁蛋白,PRO2044和Hypothetical protein根据蛋白数据库的鉴定分析结果可能是血清白蛋白的修饰产物。转铁蛋白和载脂蛋白E在以往卵泡发育机制的研究罕有报道。结论:蛋白质组学是全面分析人卵泡发育过程中卵泡液蛋白质表达变化的有效手段,筛选鉴定得到的卵母细胞成熟相关蛋白为进一步研究卵母细胞成熟机制提供了理论依据。

小鼠成熟卵丘-卵母细胞复合体蛋白质谱图的建立

目的通过聚丙烯酰胺二维凝胶电泳技术,全面展示小鼠成熟卵丘卵母细胞复合体(COC)的蛋白质表达谱。方法采用13cmpH3～10的非线性胶条进行双向凝胶电泳,分离成熟雌鼠COC总蛋白,并作质谱鉴定。结果获得了较高分辨率的小鼠成熟COC蛋白质表达谱图,并对胶上的3个蛋白点进行了质谱鉴定。结论蛋白质组学是全面分析COC蛋白表达的有效手段。成熟COC蛋白质表达谱的建立为研究卵泡发育机理提供了依据。

Electrophoresis Analysis on the Protein Expression in Heteromorphic Leaves of Populus euphratica Oliv.

[Objective] This study was to elucidate the cellular and molecular mechanism of the development of heteromorphic leaves of Populus euphratica Oliv. [Method] By employing SDS-PAGE and 2-demensional electrophoresis (2-DE) techniques,proteins in various heteromorphic leaves from the same adult tree of P. euphratica were isolated and separated to the electrophoresis technique suitable for the separation and analysis of proteins in leaves of P. euphratica tree. [Results] There were significant differences in the expressions of proteins in various heteromorphic leaves of P. euphratica tree. SDS-PAGE pattern showed that bands of proteins with molecular weight of 57.2,13.2,30.2,23.9 and 33.3 kDa were remarkably different. 2-D electrophoresis pattern presented that proteins in leaves of P. euphratica tree mainly belong to acidic proteins distributed at pH value of 5.0-6.5 and with molecular weight of 20-40 kDa; totally 73 different protein spots were observed,of which 51 were up expressed and other 22 were down expressed in the serrated ovate leaves. [Conclusion] Based on these results,we speculate that regulated gene expression in leaves of P. euphratica tree results in the generation of different shapes of leaves,in order to adapt to the surroundings better.

Diagnostic model of saliva protein finger print analysis of patients with gastric cancer

AIM:To explore the method for early diagnosis of gastric cancer by screening the expression spectrum of saliva protein in gastric cancer patients using mass spectrometry for proteomics.METHODS:Proportional peptide mass fingerprints were obtained by analysis based on proteomics matrix-assisted laser desorption ionization time-of-flight/mass spectrometry.A diagnosis model was established using weak cation exchange magnetic beads to test saliva specimens from gastric cancer patients and healthy subjects.RESULTS:Significant differences were observed in the mass to charge ratio(m/z) peaks of four proteins(1472.78 Da,2936.49 Da,6556.81 Da and 7081.17 Da) between gastric cancer patients and healthy subjects.CONCLUSION:The finger print mass spectrum of saliva protein in patients with gastric cancer can be established using gastric cancer proteomics.A diagnostic model for distinguishing protein expression mass spectra of gastric cancer from non-gastric-cancer saliva can be established according to the different expression of proteins 1472.78 Da,2936.49 Da,6556.81 Da and 7081.17 Da.The method for early diagnosis of gastric cancer is of certain value for screening special biological markers.

Applied research on serum protein fingerprints for prediction of Qi deficiency syndrome and phlegm and blood stasis in patients with non-small cell lung cancer

OBJECTIVE:This study screened serum tumor biomarkers by surface enhanced laser desorption/ionization time-of-flight mass spectrometry(SELDI-TOF-MS) to establish a subset which could be used for the prediction of Qi deficiency syndrome and phlegm and blood stasis in patients with non-small cell lung cancer;and as diagnostic model of Chinese medicine.METHODS:Serum samples from 63 lung cancer patients with Qi deficiency syndrome and phlegm and blood stasis,and 28 lung cancer patients with non-Qi deficiency syndrome and phlegm and blood stasis were analyzed using SELDI-TOF-MS with a PBS II-C protein chip reader.Protein profiles were generated using immobilized metal affinity capture(IMAC3) protein chips.Differentially-expressed proteins were screened.Protein peak clustering and classification analyses were performed using Biomarker Wizard and Biomarker Pattern software packages,respectively.RESULTS:A total of 268 effective protein peaks were detected in the 1,000-10,000 Da molecular range for the 15 serum proteins screened(P<0.05).The decision tree model was M 2284.97,with a sensitivity of 96.2% and a specificity of 66.7%.CONCLUSION:SELDI-TOF-MS techniques,combined with a decision tree model,can help identify serum proteomic biomarkers related to Qi deficiency syndrome and phlegm and blood stasis in lung cancer patients;and the predictive model can be used to discriminate between Chinese medicine diagnostic models of disease.