E.M. Santos, J.F.R. Paula, P.M.C. Motta, M.B. Heinemann, R.C. Leite, J.P.A. Haddad, H.L. Del Puerto and J.K.P. Reis
Published August 17, 2010
Genet. Mol. Res. 9 (3): 1591-1598 (2010)
DOI 10.4238/vol9-3gmr818
About the Authors
E.M. Santos, J.F.R. Paula, P.M.C. Motta, M.B. Heinemann, R.C. Leite, J.P.A. Haddad, H.L. Del Puerto and J.K.P. Reis
Corresponding author:
E.M. Santos
E-mail: elmaira27@yahoo.com.br
ABSTRACT
We compared three different protocols for DNA extraction from horse peripheral blood mononuclear cells (PBMC) and lung fragments, determining average final DNA concentration, purity, percentage of PCR amplification using β-actin, and cost. Thirty-four samples from PBMC, and 33 samples from lung fragments were submitted to DNA extraction by three different protocols. Protocol A consisted of a phenol-chloroform and isoamylic alcohol extraction, Protocol B used alkaline extraction with NaOH, and Protocol C used the DNAzol® reagent kit. Protocol A was the best option for DNA extraction from lung fragments, producing high DNA concentrations, with high sensitivity in PCR amplification (100%), followed by Protocols C and B. On the other hand, for PBMC samples, Protocol B gave the highest sensitivity in PCR amplification (100%), followed by Protocols C and A. We conclude that Protocol A should be used for PCR diagnosis from lung fragment samples, while Protocol B should be used for PBMC.
Key words: Protocol; DNA; Equine; PBMC; Lung