کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
5546463 | 1402743 | 2017 | 5 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
A comparison of DNA extraction protocols from blood spotted on FTA cards for the detection of tick-borne pathogens by Reverse Line Blot hybridization
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کلمات کلیدی
موضوعات مرتبط
علوم زیستی و بیوفناوری
علوم کشاورزی و بیولوژیک
علوم دامی و جانورشناسی
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چکیده انگلیسی
An essential step in the molecular detection of tick-borne pathogens (TBPs) in blood is the extraction of DNA. When cooled storage of blood under field conditions prior to DNA extraction in a dedicated laboratory is not possible, the storage of blood on filter paper forms a promising alternative. We evaluated six DNA extraction methods from blood spotted on FTA Classic® cards (FTA cards), to determine the optimal protocol for the subsequent molecular detection of TBPs by PCR and the Reverse Line Blot hybridization assay (RLB). Ten-fold serial dilutions of bovine blood infected with Babesia bovis, Theileria mutans, Anaplasma marginale or Ehrlichia ruminantium were made by dilution with uninfected blood and spotted on FTA cards. Subsequently, DNA was extracted from FTA cards using six different DNA extraction protocols. DNA was also isolated from whole blood dilutions using a commercial kit. PCR/RLB results showed that washing of 3Â mm discs punched from FTA cards with FTA purification reagent followed by DNA extraction using Chelex® resin was the most sensitive procedure. The detection limit could be improved when more discs were used as starting material for the DNA extraction, whereby the use of sixteen 3Â mm discs proved to be most practical. The presented best practice method for the extraction of DNA from blood spotted on FTA cards will facilitate epidemiological studies on TBPs. It may be particularly useful for field studies where a cold chain is absent.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Ticks and Tick-borne Diseases - Volume 8, Issue 1, January 2017, Pages 185-189
Journal: Ticks and Tick-borne Diseases - Volume 8, Issue 1, January 2017, Pages 185-189
نویسندگان
Zerihun Hailemariam, Jabbar Sabir Ahmed, Peter-Henning Clausen, Ard Menzo Nijhof,