DNA Fingerprinting of brinjal (Solanum melongena L) varieties and related species
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Date
2013
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Centre for Plant Biotechnology and Molecular Biology, College of Horticulture, Vellanikkara
Abstract
Brinjal (Solanum melongena L.) also known as aubergine or eggplant, is a
member of the family Solanaceae. It is an important vegetable in central, southern
and south-east Asia, and in a number of African countries. In India, it is one of the
most common, popular and principal vegetable crop grown throughout the country
except in higher altitudes.
The study entitled “DNA fingerprinting of brinjal (Solanum melongena L.)
variet ies and related species” was carried out at the Center for Plant Biotechnology
and Molecular Biology, College of Horticulture Vellanikkara during the period 20112013.
The objective of study was to characterize the four brinjal varieties released by Kerala Agricultural University using molecular markers like ISSR and SSR and to
develop a DNA fingerprint specific to each variety. Three superior accessions and
two wild relatives were also characterized to detect the level of variability.
The brinjal varieties/accessions selected include three Solanum melongena
varieties- Surya, Swetha, Haritha and the hybrid Neelima; three accessions- SM 116,
SM 396, SM 397 and the two wild relatives S. melongena var. insanum and S.
macrocarpon. Breeder seeds were obtained from the Department of Olericulture,
College of Horticulture, Kerala Agricultural University and maintained at CPBMB,
COH and further used for the study.
Morphological characters were recorded for six vegetative and five
reproductive characters according to the descriptor of NBPGR. The data recorded
was used to validate the genotypes which was used for fingerprinting. DNA
extraction was done according to the method described by Rogers and Bendich 1994.
The RNA contamination was completely removed through RNase treatment. Good
quality DNA with UV absorbance ratio (OD
260
/OD
) 1.80 – 1.89 was used for
further analysis.
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Thirty six ISSR primers and sixty one SSR primer pairs were screened with
DNA of brinjal genotypes for amplification and those which gave reliable distinct
banding pattern were selected for further amplification/fingerprinting. The PCR product obtained from ISSR and SSR analysis were separated on 2 per cent agarose
gel and amplification patterns recorded.
The genomic DNA from each genotype was amplified with 10 each of
selected ISSR and SSR primers. The amplification pattern was scored and depicted
to develop fingerprint for each brinjal genotype.
The Resolving power (Rp) of the ISSR primers was calculated and the values
ranged between 9.9 and 28.44 indicating the capacity of the primers selected to
distinguish the genotype. The Polymorphic Information Content (PIC) was also
calculated and it ranged between 0.83 and 0.96 for ISSR primers, further indicating
the suitability of primers to detect polymorphism. The PIC value for SSR primers
were zero and not suitable to detect polymorphism.
Distinct amplicons developed through ISSR and SSR analysis were used to
develop the DNA fingerprint of brinjal genotypes. Sharing of amplicons for each
primer with other genotypes was also analyzed and demarcated with different colour
codes in the fingerprint developed. Most of the amplicons were found shared among
the genotypes indicating their genetic uniformity. However, the pattern of sharing
was different and good enough to separate out the varieties and distinct amplicons
were observed in the genotypes.
The ISSR and SSR banding pattern was scored and analysed for their
uniformity/variability using the software NTSYS pc (version 2.02i). Similarity
coefficient ranged from 0.51 to 0.84. The highest similarity (84%) was observed
between the brinjal hybrid Neelima and its maternal parent Surya. The S.
macrocarpon was the most distinct one from other brinjal genotypes with 49 per cent
variability.
Separate fingerprint were developed for all the four varieties, three accessions
and two wild relatives through ISSR and SSR analysis. The DNA fingerprints thus
developed could be utilised for the variety registration and settling IPR issues.
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