从粪便标本中快速可靠地提取原生动物寄生虫DNA。
Fast and reliable extraction of protozoan parasite DNA from fecal specimens.
作者信息
da Silva A J, Bornay-Llinares F J, Moura I N, Slemenda S B, Tuttle J L, Pieniazek N J
机构信息
Division of Parasitic Diseases, National Center for Infectious Diseases, Centers for Disease Control and Prevention, Public Health Service, U.S. Department of Health and Human Services, Atlanta, Georgia 30341-3724, USA.
出版信息
Mol Diagn. 1999 Mar;4(1):57-64. doi: 10.1016/s1084-8592(99)80050-2.
BACKGROUND
Polymerase chain reaction (PCR) detection of intestinal protozoa in fecal specimens is hampered by poor recovery of DNA and by the presence of PCR inhibitors. In this study we describe a novel method for DNA extraction from such specimens containing spores and oocysts of Enterocytozoon bieneusi and Cryptosporidium parvum, respectively.
METHODS AND RESULTS
Extraction was done using commercial kits modified to maximize the recovery and purity of extracted DNA. In comparison with a procedure we previously reported, we estimate that this method may increase the sensitivity of parasite DNA detection in fecal specimens up to tenfold. An additional advantage of this method is that up to 12 samples may be processed simultaneously within 2 hours.
CONCLUSIONS
By using this method, we were able to increase reproducibility of PCR amplification on fecal specimens and significantly reduce the hands-on time required to process the samples.
背景
粪便标本中肠道原生动物的聚合酶链反应（PCR）检测受到DNA回收率低和PCR抑制剂存在的阻碍。在本研究中，我们描述了一种从此类分别含有微小隐孢子虫和贝氏等孢球虫孢子和卵囊的标本中提取DNA的新方法。
方法与结果
使用经过改良的商业试剂盒进行提取，以最大限度提高提取DNA的回收率和纯度。与我们之前报道的方法相比，我们估计该方法可将粪便标本中寄生虫DNA检测的灵敏度提高至原来的十倍。该方法的另一个优点是，可在2小时内同时处理多达12个样本。
结论
通过使用该方法，我们能够提高粪便标本PCR扩增的重现性，并显著减少处理样本所需的实际操作时间。
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