Isolation of genomic DNA is a prerequisite for assessment of milk quality. As a source of genomic DNA, milk somatic cells from milking ruminants are practical, animal friendly, and cost-effective sources. Extracting DNA from milk can avoid the stress response caused by blood and tissue sampling of cows. In this study, we optimized a novel DNA extraction method for amplifying long (>1,000 bp) DNA fragments and used it to evaluate the isolation of DNA from small amounts of milk. The techniques used for the separation of milk somatic cell were explored and combined with a sodium dodecyl sulfate (SDS)-phenol method for optimizing DNA extraction from milk. Spectrophotometry was used to determine the concentration and purity of the extracted DNA. Gel electrophoresis and DNA amplification technologies were used for to determine DNA size and quality. The DNA of 112 cows was obtained from milk (samples of 13 ± 1 mL) and the corresponding optical density ratios at 260:280 nm were between 1.65 and 1.75. Concentrations were between 12 and 45 μg/μL and DNA size and quality were acceptable. The specific PCR amplification of 1,019- and 729-bp bovine DNA fragments was successfully carried out. This novel method can be used as a practical, fast, and economical mean for long genomic DNA extraction from a small amount of milk.
The extraction of high-quality DNA from processed dairy products is often the crucial step in an authentication process by PCR-based methods. In this study, we optimized a novel DNA extraction method for milk powder and used the extracted DNA for identification of milk powder based on PCR analysis. The DNA quality was assessed by amplifying target sequences from mitochondrial genes, as well as by monitoring the yield, purity, and integrity of the extracted DNA. In addition, a laboratory adulteration model of milk powder was detected by PCR-based methods (PCR and real-time PCR) using primers targeting the mitochondrial 12S rRNA gene. Results showed that a sufficient amount and quality of DNA could be isolated from milk powder with this method. Both PCR and real-time PCR detection of cow milk compositions in goat milk powder further confirmed the DNA extracted with this extraction method could be widely used in addressing milk powder adulterant by a PCR-based method.
Relationships between viscosity and the contents of macromolecular substances (milk protein, casein, whey protein and milk fat) from raw milk were studied at the temperature of 2-5 o C (a storage temperature in a household refrigerator), 10 o C and 20 o C (a near-room temperature) during milk storage. An approach by means of mathematical models was used in this study to analyze the relationships between the viscosity and the contents of these macromolecular substances from milk at the three temperatures. The results indicated that the milk viscosity initially remained quite steady for a period of time, followed by an increase for a short-period, then a decrease during the subsequent days, and had a sharp rise in the final days at the temperatures of 10 o C and 20 o C, while the viscosity of milk stored at the temperature of 2-5 o C presented a decrease initially and a sharp rise in the final days. Significant correlations were also found between the value of viscosity and the contents of macromolecular substances along with storage time extended at the three temperatures. The correlation models based on the viscosity and the contents of macromolecular substances had good fit with high correlation coefficients (R 2 >0.8000) and could well explain the relevance of viscosity and the contents of macromolecular substances at a significance level that was less than or equal to 0.0067. Meanwhile, these correlation models contributed to a better understanding of the change of milk system during storage in view of providing tools to develop high quality dairy products with desirable characteristics, and could also be useful in process design and control, as well as quality control and real-time monitoring of deterioration of the raw milk.
This study investigated the expression of novel mutations of the β2-microglobulin (B2M) gene in bovine genomic DNA isolated from milk-borne leukocytes and tested the association of these mutations with milk quality traits in Holstein cows. DNA was extracted using a novel protocol allowing for the amplification of long fragments of DNA isolated from small amounts of unprocessed milk samples. Genomic mutations at the B2M locus were explored by DNA sequencing using specific primers and a cluster of five identified within a 729-bp region of intron 3. Specifically, mutations at 7008(G>T), 7202(A>G), 7432(G>T) and 7437(G>C) were all found to be significantly associated with milk protein percentage (P < 0.05), and a mutation at 7026(G>A) was significantly associated with somatic cell count (P < 0.05). The potential to use these mutations as further tools for the pre-evaluation of milk quality is discussed. Valoración de la calidad de la leche utilizando novedosas mutaciones del gen B2M en el ADN bovino obtenido de la leche RESUMEN El presente estudio investigó cómo se manifiestan mutaciones novedosas del gen β2-microglobulina en el ADN genómico bovino aislado de leucocitos transmitidos en la leche, a la vez que sometió a prueba la asociación de dichas mutaciones con las características de calidad de la leche obtenida de vacas Holstein. El ADN se extrajo utilizando un protocolo innovador que permite amplificar largos fragmentos del ADN aislados de pequeñas cantidades de muestras de leche sin procesar. Se examinaron las mutaciones genómicas ocurridas en el locus del (B2M) mediante la secuenciación del ADN, utilizando para ello partidores específicos y un clúster de cinco, identificados en una región de 729 bp del intrón 3. Específicamente, se determinó que las mutaciones en 7008 (G>T), 7202 (A>G), 7432 (G>T) y 7437 (G>C) se encuentran significativamente asociadas con el porcentaje de proteína láctea (MPP) (P<0,05), y que una mutación en 7026 (G>A) se asocia de manera significativa con el conteo de células somáticas (SCC) (P<0,05). Para finalizar, se discute la posibilidad de usar estas mutaciones como herramientas adicionales para la preevaluación de la calidad de la leche.
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