African Journal of Biotechnology Vol. 6 (3), pp. 184-187, 5 February 2007   

ISSN 1684–5315 © 2007 Academic Journals        

Full Length Research Paper

Influence of EDTA and magnesium on DNA extraction from blood samples and specificity of polymerase chain reaction

Khosravinia, H.^1* and Ramesha, K. P.^2y

¹Department of Technology of Animal Products, Agriculture Faculty, Lorestan University, P.B. 465, Khoramabad-68135, Lorestan, Iran.

²Department of Biochemistry, Indian Institute of Science, Bangalore-560012, India.

*Corresponding author. E-mail: Khosravi_fafa@yahoo.com. Ph:  (+98)(+661) 3203714 (Off.), (+98)(+661) 2212851 (Res.), 09166673705 (Mob.). Fax: (+98)(+661) 4200289.

^yPresent Address: NRC on Yak, Dirang-790101, Via Bomdila, Arunachal Pradesh, India.

Accepted 8 December, 2006

This study consisting of two trails conducted to examine the impact of initial EDTA level added to blood samples on quantity and quality of genomic DNA isolated from avian fresh blood and the influence of initial EDTA level with various levels of MgCl₂ added to polymerase chain reaction (PCR) final volume on amplification pattern. EDTA level added to collected blood samples had no significant impact on quantity as well as quality of extracted genomic DNA. However, higher levels of EDTA increased the OD₂₆₀ and enhanced the OD₂₆₀/OD₂₈₀ ratio beyond 1.8-1.9 which is broadly accepted as an indicator of high quality DNA. To avoid such an error, EDTA level in initial blood sample must not exceed 9 µg/µl blood. The initial amount of EDTA has no influence on PCR process if a valid DNA isolation protocol is used. Addition of MgCl₂ from 1.0 to 2.4 µl in a final volume of 25 µl could support the amplification properly. Low levels of MgCl₂ results in incomplete amplification but levels higher than 2.4 µl impedes the amplification by negative interference to the whole reactions.

Key words: EDTA, DNA extraction, Mg²⁺ concentration, PCR.