[1]王洁敏,向 芃,顾乐怡.不同预处理方法对提取尿液脱落细胞mRNA质量的影响[J].现代检验医学杂志,2016,31(04):125-127，130.[doi:10.3969/j.issn.1671-7414.2016.04.035]
　WANG Jie-min,XIANG Peng,GU Le-yi.Effects of Different Pretreatment Methods on mRNA Quality and Quantity of Exfoliated Urinary Cells[J].Journal of Modern Laboratory Medicine,2016,31(04):125-127，130.[doi:10.3969/j.issn.1671-7414.2016.04.035]
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不同预处理方法对提取尿液脱落细胞mRNA质量的影响()
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《现代检验医学杂志》[ISSN:/CN:]

卷:
第31卷
期数:
2016年04期
页码:
125-127，130
栏目:
研究简报·实验技术
出版日期:
2016-08-10

文章信息/Info

Title:
Effects of Different Pretreatment Methods on mRNA Quality and Quantity of Exfoliated Urinary Cells
文章编号:
1671-7414(2016)04-125-04
作者:
王洁敏; 向 芃; 顾乐怡
上海交通大学医学院附属仁济医院肾脏科,上海 200127
Author(s):
WANG Jie-min; XIANG Peng; GU Le-yi
Renal Division, Renji Hospital,Shanghai Jiaotong University School of Medicine,Shanghai200127,China
关键词:
慢性肾脏病;  尿液脱落细胞;  mRNA;  足细胞
分类号:
R446.12
DOI:
10.3969/j.issn.1671-7414.2016.04.035
文献标志码:
A
摘要:
目的 探索合适的处理尿标本的方法获得尿脱落细胞mRNA。方法 比较尿液容量、尿液标本冻存时间、处理方式对提取尿液脱落细胞RNA的产量与质量的影响,并进行qPCR的试验。结果 40 ml尿液标本组及10 ml尿液标本组RNA总量均值分别为886.94±222.93 ng及211.22±62.01 ng,两组A260nm/A280nm比值分别为1.80±0.10和1.77±0.09; 尿液标本量与RNA总量相关(t=3.604,P=0.002),不同尿液标本量RNA质量差异无统计学意义(t=0.708,P值>0.05)。实时离心提取及冰冻保存尿液7天后提取两种预处理方法RNA总量分别为886.94±945.84 ng及881.50±829.02 ng,A260nm/A280nm比值分别为1.80±0.10及1.80±0.87; RNA总量与A260nm/A280nm比值差异无统计学意义(t=-0.221～0.085,P值均>0.05)。直接冰冻尿液标本与TRIzol保存沉渣细胞两种预处理方法RNA总量分别为637.81±525.24 ng及639.86±535.42 ng,A260nm/A280nm比值分别为1.84±0.13及1.83±0.96; RNA总量与A260nm/A280nm比值差异无统计学意义(t=-0.47～0.293,P值均>0.05)。尿脱落细胞mRNA中可以检测到足细胞的标记蛋白WT-1,podocin和synaptopodin的基因表达。结论 尿液标本量是提取脱落细胞RNA量的关键因素,-80℃低温冻存尿标本7天或Trizol预处理冻存均对提取尿液沉渣细胞RNA数量和质量无影响。
Abstract:
Objective To explore a proper method to obtainexfoliated cells mRNA for detection in urine specimen.Methods Urine volume,different pretreating ways and frozen storage time on urinary sample were used to collect mRNA.A260nm/A280nm and A260nm/A280nm ratio,as well as qPCR were performed.Results Amount of mRNA was determinded by urine volume,but neitherA260nm/A280nm or A260nm/A280nm ratio wasrealated with urine volume.Compared with real-time extraction process,freezing urine samples for 7 days showed no statistical difference(P>0.05)in total RNA and A260nm/A280nm or A260nm/A280nm ratio.There was no significant difference in total RNA and A260nm/A280nm or A260nm/A280nm ratio between direct freezing urine samples and Trizol-treated urine exfoliated cells.Podocyte marker including WT-1,podocin and synaptopodin mRNAs were detected in urine sediment cells.Conclusion Quantity of mRNA was determinded by urine volume.Directly freezing urinesamples at -80 ℃ for 7 days or Trizol-treating urine sediment cells have no effect on lowing quality and quantity of urinary sediment mRNA.

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备注/Memo

备注/Memo:
基金项目:国家自然科学基金资助项目(NO.81270781)。
作者简介:王洁敏(1989-),女,在读研究生,研究方向:慢性肾脏病。
通讯作者: 顾乐怡(1973-),男,肾脏内科医师,E-mail:guleyi@aliyun.com。
更新日期/Last Update: 2016-08-10