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Short Communication
DNA extraction from ghee and beef species identification using polymerase
chain reaction (PCR) assay
Nooratiny, I., 3,4Sahilah, A.M., 2Alif Alfie, A. R. and 5Mohd. Farouk, M. Y.
1,2*
Article history
Abstract
A technique to isolate DNA from ghee was developed for the authentication of beef fat product.
The method was based on pre-mixed ghee with phosphate buffer solution (PBS) prior to DNA
extraction using Epicentre extraction method. The recovery of beef DNA was then analysed
by polymerase chain reaction (PCR) using beef species-specific oligonucleotide primers which
targeted the mitochondria DNA (mtDNA) of cytochrome b (cyt b) gene. The amplicon was 274
bp in size. The developed ghee extraction method offers a high yield of DNA providing 100 ng
per l and useful for validating beef fat product.
Keywords
Ghee
Beef
CYTB gene
SIM B and Polymerase
Chains Reaction (PCR)
Introduction
The DNA-based approaches of polymerase
chain reaction (PCR) techniques are well known as
a specific, reproducible, sensitive, rapid processing
time and low costs. However, the successful of this
approach is limited by the presence of inhibitors in
food matrix. The recovery of good quality DNAis
vital in the PCR process. Without DNA or poor
quality of DNA, the oligonucleotide primers are not
able to amplify the targeted gene in the PCR assay.
In meat specification, either nuclear or
mitochondrial DNA (nDNA and mtDNA) genes have
been targeted in PCR analysis. Nuclear DNA is larger
molecule arranged into chromosomes and contains a
greater variation in its type of sequences compared
to mtDNA. The mtDNA is commonly used for
species identification in food analysis (Meyer et al.,
1994; Matsunaga et al., 1999; Che Man et al., 2007;
Sahilah et al., 2011) especially cytochrome b (cyt b)
gene. High copy number of mtDNA is found in the
cells, it remain intact during food processing thereby
minimizing DNA degradation and does not contain
any introns (Unseld et al., 1995). The major problem
in DNA extraction from ghee is due to high fats
content that may inhibit PCR analysis. The inhibitors
in food can be in the form of polysaccharides, protein,
humic acids and others. Some of the inhibitors are
*Corresponding author.
Email: tinytoon79@yahoo.com
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