Optimization of RNA yield, purity and mRNA copy number by treatment of urine cell pellets with RNAlater
Optimization of RNA yield, purity and mRNA copy number by treatment of urine cell pellets with RNAlater
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DOI：
10.1016/s0022-1759(03)00237-0
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发表时间：
2003-08-01
影响因子：
2.2
通讯作者：
Suthanthiran, M
中科院分区：
文献类型：
--
作者：
Medeiros, M;Sharma, VK;Suthanthiran, M
背景：我们已经表明，测量尿细胞中细胞毒性攻击蛋白和颗粒酶B的mRNA是诊断人类同种异体肾移植急性排斥反应的一种无创手段。尿细胞mRNA研究已经在其他患者群体（如癌症患者）中获得了有用的信息。然而，从尿细胞中分离出足够和高质量的核糖核酸（RNA）是有问题的。RNAlater是一种RNA稳定溶液，据报道可以优化室温保存的肿瘤组织和富含色素的眼部组织中RNA的分离。方法：探讨在尿细胞微球中加入RNAlater是否能提高RNA产量、提高纯度并便于检测低丰度mrna。我们使用实时定量聚合酶链反应（PCR）检测，检测了同种异体肾移植受者尿液标本和肾活检组织中组成表达基因18S rRNA和颗粒酶B和转化生长因子- β (1) mRNA的表达水平。结果：RNA产率(P
Background: We have shown that measurement of mRNA for cytotoxic attack proteins perform and granzyme B in urinary cells is a noninvasive means of diagnosing acute rejection of human renal allografts. Urinary cell mRNA studies have yielded useful information in other patient populations such as patients with cancer. The isolation of sufficient and high quality ribonucleic acid (RNA) from urinary cells however is problematic. RNAlater, an RNA stabilization solution, has been reported to optimize RNA isolation from tumor tissues stored at room temperature and from pigment-rich ocular tissues. Methods: We explored whether the addition of RNAlater to urine cell pellets improves RNA yield, enhances purity and facilitates measurement of low abundance mRNAs. We measured, with the use of real-time quantitative polymerase chain reaction (PCR) assay, levels of expression of a constitutively expressed gene 18S rRNA and mRNA for granzyme B and transforming growth factor-beta(1) (TGF-beta(1)) in urine specimens and renal biopsies obtained from renal allograft recipients. Results: RNA yield (P