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CLC number: Q503

On-line Access: 2012-04-06

Received: 2011-06-18

Revision Accepted: 2011-09-29

Crosschecked: 2012-02-29

Cited: 14

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Journal of Zhejiang University SCIENCE B 2012 Vol.13 No.4 P.239-243

http://doi.org/10.1631/jzus.B1100194


Extraction of DNA suitable for PCR applications from mature leaves of Mangifera indica L.


Author(s):  Muhammad Abubakkar Azmat, Iqrar Ahmad Khan, Hafiza Masooma Naseer Cheema, Ishtiaq Ahmad Rajwana, Ahmad Sattar Khan, Asif Ali Khan

Affiliation(s):  Centre of Agricultural Biochemistry and Biotechnology, University of Agriculture, Faisalabad, Pakistan; more

Corresponding email(s):   knnike@yahoo.com

Key Words:  Extraction buffer, Mango, Polyphenols, DNA isolation, Simple sequence repeat (SSR), Secondary metabolites


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Muhammad Abubakkar Azmat, Iqrar Ahmad Khan, Hafiza Masooma Naseer Cheema, Ishtiaq Ahmad Rajwana, Ahmad Sattar Khan, Asif Ali Khan. Extraction of DNA suitable for PCR applications from mature leaves of Mangifera indica L.[J]. Journal of Zhejiang University Science B, 2012, 13(4): 239-243.

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author="Muhammad Abubakkar Azmat, Iqrar Ahmad Khan, Hafiza Masooma Naseer Cheema, Ishtiaq Ahmad Rajwana, Ahmad Sattar Khan, Asif Ali Khan",
journal="Journal of Zhejiang University Science B",
volume="13",
number="4",
pages="239-243",
year="2012",
publisher="Zhejiang University Press & Springer",
doi="10.1631/jzus.B1100194"
}

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%A Muhammad Abubakkar Azmat
%A Iqrar Ahmad Khan
%A Hafiza Masooma Naseer Cheema
%A Ishtiaq Ahmad Rajwana
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%A Asif Ali Khan
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%I Zhejiang University Press & Springer
%DOI 10.1631/jzus.B1100194

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A1 - Hafiza Masooma Naseer Cheema
A1 - Ishtiaq Ahmad Rajwana
A1 - Ahmad Sattar Khan
A1 - Asif Ali Khan
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DOI - 10.1631/jzus.B1100194


Abstract: 
Good quality deoxyribonucleic acid (DNA) is the pre-requisite for its downstream applications. The presence of high concentrations of polysaccharides, polyphenols, proteins, and other secondary metabolites in mango leaves poses problem in getting good quality DNA fit for polymerase chain reaction (PCR) applications. The problem is exacerbated when DNA is extracted from mature mango leaves. A reliable and modified protocol based on the cetyltrimethylammonium bromide (CTAB) method for DNA extraction from mature mango leaves is described here. High concentrations of inert salt were used to remove polysaccharides; Polyvinylpyrrolidone (PVP) and β-mercaptoethanol were employed to manage phenolic compounds. Extended chloroform-isoamyl alcohol treatment followed by RNase treatment yielded 950‒1050 µg of good quality DNA, free of protein and RNA. The problems of DNA degradation, contamination, and low yield due to irreversible binding of phenolic compounds and coprecipitation of polysaccharides with DNA were avoided by this method. The DNA isolated by the modified method showed good PCR amplification using simple sequence repeat (SSR) primers. This modified protocol can also be used to extract DNA from other woody plants having similar problems.

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Open peer comments: Debate/Discuss/Question/Opinion

<1>

Mudassir Iqbal@AARI<asuaf@gmail.com>

2012-01-25 18:10:05

This is an excellent protocol i tried this protocol to extract DNA from mature leaves of mulberry. The DNA obtained was of very good quality.

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