Resumen

SANCHO-BLANCO, Carolina; JIMENEZ-ALFARO, Esteban J.; MOLINA-BRAVO, Ramón  y  UMANA-CASTRO, Rodolfo. Incubation, pre-lysis and post-purification on the yield and purity of nucleic acids extracted from blood of domestic goats contained in FTA cards. Rev. mex. de cienc. pecuarias [online]. 2022, vol.13, n.1, pp.311-322.  Epub 06-Jun-2022. ISSN 2448-6698.  https://doi.org/10.22319/rmcp.v13i1.5890.

Molecular techniques require extractions of nucleic acids in adequate quantity and purity. This work describes a generalized linear model (GLM) of an adjusted factor with fixed effects on nucleic acid yield (ng/μl) and purity (A260/A280 and A260/A230), for five methods of DNA extraction using FTA cards with goat (Capra aegagrus hircus) blood. Two commercial methods based on silica columns (Invitrogen and Macherey Nagel; MN), the chelating resin method (Chelex), the CTAB method and the phenol-chloroform-isoamyl alcohol (PCI) method were tested. Additionally, for MN, an incubation step with PBS (Phosphate Buffered Saline) buffer at high temperature prior to lysis and a purification step post extraction were evaluated using a fixed-effect model of two factors with interaction. DNA concentrations and purity ratios were variable; the highest concentration was obtained with the MN kit (170.45 ng/μl), but with deficiencies in purity (0.32 of A260/A230, 0.34 of A260/A280). Despite this, all extraction methods generated PCR products with specific D-loop primers (mtDNA). The combined effect of the pre-incubation and post-purification stages yielded satisfactory purity values (1.89 for A260/A230 and 1.65 for A260/A280), as well as concentration ratios (476.78 ng/μl) with low variability. In conclusion, the concentration and purity of DNA from blood samples is greatly improved when using a commercial kit in combination with pre-lysis incubation and post-extraction purification. These nucleic acids are suggested for use in potential molecular applications a posteriori.

Palabras llave : DNA; Silica columns; CTAB; Phenol-chloroform; Chelex; PCR; Small ruminants.