Molecular identification of Taenia specimens after long-term preservation in formalin
Molecular identification of Taenia specimens after long-term preservation in formalin
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DOI：
10.1016/j.parint.2010.12.001
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发表时间：
2011-06-01
影响因子：
1.9
通讯作者：
Eom, Keeseon S.
中科院分区：
文献类型：
--
作者：
Jeon, Hyeong-Kyu;Kim, Kyu-Heon;Eom, Keeseon S.
博物馆收藏的带绦虫标本，大部分通常以福尔马林固定剂永久保存，主要用作形态学检查。本研究的目的是提高带绦虫鉴定，即使是保存在福尔马林最多81年。带绦虫由瑞士自然历史博物馆的寄生虫收集单位以及印度尼西亚、日本和韩国的单位收集。将少量福尔马林固定的组织（100 mg）在液氮中粉碎，然后在Tris-EDTA缓冲液中浸泡3 - 5 h。然后将样品在SDS和蛋白酶K（20 mg/ml）中于56 ℃消化3 - 5 h。加入蛋白酶K（20 mg/ml）、SDS和十六烷基三甲基溴化铵（CTAB）后，在65 ℃继续孵育3小时。从这个额外的步骤中获得了最大的基因组DNA产量，并且用这种提取方法获得的基因组DNA的质量似乎与在福尔马林固定剂中的储存时间的持续时间无关。应用PCR和cox1基因80-428位DNA序列对带绦虫进行分子鉴定。t.在印度尼西亚、日本和韩国的分离物中检测到亚洲丝核菌。从福尔马林固定的博物馆收藏品中提取基因组DNA方法的改进将有助于寄生虫的分子鉴定。(C)2010爱思唯尔爱尔兰有限公司版权所有。
The majority of Taenia tapeworm specimens in the museum collections are usually kept in a formalin fixative for permanent preservation mainly for use in morphological examinations. This study aims to improve Taenia tapeworm identification even of one preserved in formalin for a maximum of 81 years. Taenia tapeworms were collected by the parasite collection unit of the Swiss Natural History Museum and from units in Indonesia, Japan and Korea. A small amount of formalin-fixed tissue (100 mg) was crushed in liquid nitrogen and then soaked in a Tris-EDTA buffer for 3-5 h. The sample was then digested in SDS and proteinase K (20 mg/ml) for 3-5 h at 56 degrees C. After the addition of proteinase K (20 mg/ml), SDS and hexadecyl-trimethyl-ammonium bromide (CTAB), incubation was continued for another 3 h at 65 degrees C. A maximum yield of genomic DNA was obtained from this additional step and the quality of genomic DNA obtained with this extraction method seemed to be independent of the duration of storage time in the formalin fixative. The molecular identification of Taenia tapeworms was performed by using PCR and DNA sequences corresponding to position 80-428 of cox1 gene. T. asiatica was detected in the isolates of Indonesia, Japan and Korea. Improvements in the genomic DNA extraction method from formalin fixed museum collections will help in the molecular identification of parasites. (C) 2010 Elsevier Ireland Ltd. All rights reserved.