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گواهی نمایه سازی مقاله Comparison Two Methods of Extraction RNA Of liver Cow

عنوان مقاله: Comparison Two Methods of Extraction RNA Of liver Cow
شناسه (COI) مقاله: NIAC01_043
منتشر شده در اولین کنفرانس بین المللی ایده های نو در کشاورزی در سال ۱۳۹۲
مشخصات نویسندگان مقاله:

Seyed Mohammad Reza Motafeghi - Department of Animal Science, Young Researchers Club, Khorasgan (Isfahan) Branch, Islamic Azad University, Isfahan, Iran.
Ahmad Mahmoudi - Department of Animal Science, Young Researchers Club, Khorasgan (Isfahan) Branch, Islamic Azad University, Isfahan, Iran.
Maryam Shahriar - Department of Entomology, Young Researchers Club, Khorasgan (Isfahan) Branch, Islamic Azad University, Isfahan, Iran

خلاصه مقاله:
Since the preparation of purified and high-quality RNA is one of the most essential elements in the preparation of cDNA, low-quality extracted RNA can result in improper subsequent analysis. Due to the differences in structure of DNA and RNA, RNA molecule is more sensitive and can be easily decomposed. On the other hands, RNA degradation enzyme, i.e. RNase is abundantly available and broad range and can often act without the need of co-factor. Thus, all the stages through RNA extraction procedure must be controlled and RNase-free devices should be utilized. Given the major role of quantity and quality of extracted RNA on downstream analysis, the present study aimed to compare manual and column-based method for RNA extraction.Methods: cow’s liver was used as the RNA origin tissue. Samples were homogenized in nitrogen and the RNA extraction was carried out using both manual (RNX plus solution) and column-based method (Hybrid-R, GeneAll, South Korea). In order to scrutiny of the RNA quality, the ratio absorbance of 260:280 (nm:nm) was measured. Impurities such as proteins, phenol and other soluble components in the sample solution which mimic the absorbance ratio, agarose gel electrophoresis was carried out. In addition, the RNAs were treated with DNase enzyme to make sure the DNA-free extraction. Thereafter, the obtained RNA by the both methods underwent cDNA synthesis using Oligo dT primers followed by PCR by β-Actin primers.Results: The results showed that using Hybrid-R kit resulted in better RNA extraction in both qualitatively and quantitatively. However, after PCR-reaction, similar results were observed for the both extraction methods. This may be due to this fact that end-point PCR machine can amplify different quantity of amplicons because of large number of cycles, 35 cycles in thisexperiment. Nevertheless, extraction by the column-based method was more reliable without further requirement of DNase treatment.

کلمات کلیدی:
RNA extraction, sheep tissues, β-Actin, RT-PCR

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