| Autor: |
Alemán-Duarte MI; Laboratorio de Microbiología Industrial, Centro Universitario de Ciencias Exactas e Ingenierías, Universidad de Guadalajara, Blvd, Gral, Marcelino García Barragán 1421, Olímpica, Guadalajara 44430, Mexico., Aguilar-Uscanga BR; Laboratorio de Microbiología Industrial, Centro Universitario de Ciencias Exactas e Ingenierías, Universidad de Guadalajara, Blvd, Gral, Marcelino García Barragán 1421, Olímpica, Guadalajara 44430, Mexico., García-Robles G; Laboratorio de Microbiología Industrial, Centro Universitario de Ciencias Exactas e Ingenierías, Universidad de Guadalajara, Blvd, Gral, Marcelino García Barragán 1421, Olímpica, Guadalajara 44430, Mexico., Ramírez-Salazar FJ; Laboratorio de Microbiología Industrial, Centro Universitario de Ciencias Exactas e Ingenierías, Universidad de Guadalajara, Blvd, Gral, Marcelino García Barragán 1421, Olímpica, Guadalajara 44430, Mexico., Benítez-García I; Unidad Académica de Ingeniería en Biotecnología, Universidad Politécnica de Sinaloa (UPSIN), Carretera Municipal Libre Mazatlán Higueras Km 3 Col. Genaro Estrada, Mazatlán 82199, Mexico., Balcázar-López E; Laboratorio de Microbiología Industrial, Centro Universitario de Ciencias Exactas e Ingenierías, Universidad de Guadalajara, Blvd, Gral, Marcelino García Barragán 1421, Olímpica, Guadalajara 44430, Mexico., Solís-Pacheco JR; Laboratorio de Microbiología Industrial, Centro Universitario de Ciencias Exactas e Ingenierías, Universidad de Guadalajara, Blvd, Gral, Marcelino García Barragán 1421, Olímpica, Guadalajara 44430, Mexico. |
| Abstrakt: |
The human milk microbiota (HMM) of healthy women can vary substantially, as demonstrated by recent advances in DNA sequencing technology. However, the method used to extract genomic DNA (gDNA) from these samples may impact the observed variations and potentially bias the microbiological reconstruction. Therefore, it is important to use a DNA extraction method that is able to effectively isolate gDNA from a diverse range of microorganisms. In this study, we improved and compared a DNA extraction method for gDNA isolation from human milk (HM) samples to commercial and standard protocols. We evaluated the extracted gDNA using spectrophotometric measurements, gel electrophoresis, and PCR amplifications to assess its quantity, quality, and amplifiability. Additionally, we tested the improved method's ability to isolate amplifiable gDNA from fungi, Gram-positive and Gram-negative bacteria to validate its potential for reconstructing microbiological profiles. The improved DNA extraction method resulted in a higher quality and quantity of the extracted gDNA compared to the commercial and standard protocols and allowed for polymerase chain reaction (PCR) amplification of the V3-V4 regions of the 16S ribosomal gene in all the samples and the ITS-1 region of the fungal 18S ribosomal gene in 95% of the samples. These results suggest that the improved DNA extraction method demonstrates better performance for gDNA extraction from complex samples such as HM. |
