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Variability Among Selective Guava (psidium Guajava L.) Varieties Revealed by Morphology and Rapd Marker

Farzana Alam
Department of Genetic Engineering and Biotechnology, Jagannath University, Dhaka-1100, Bangladesh

Kazi Didarul Islam
Department of Genetic Engineering and Biotechnology, Jagannath University, Dhaka-1100, Bangladesh

S. M. Mahbubur Rahman
Department of Genetic Engineering and Biotechnology, Jagannath University, Dhaka-1100, Bangladesh

Abstract/ Executive Summary:

The research was conducted for the assessment of genetic diversity using both morphological and random amplified polymorphic DNA (RAPD) analysis of twelve guava (Psidium guajava L.) varieties growing in Bangladesh. Morphological characterization of guava varieties showed a wide range of variation. The highest variability was observed between Poly and Jelly varieties.Polymerase chain reaction with 5 arbitrary 10-mer and 3 arbitrary 12- mer RAPD primers produced a total of 50 bands of which 75.23 percent were polymorphic. The highest percentage of polymorphic loci (100%) was observed for primer A and the lowest (50%) for A03 primer. The UPGMA dendrogram revealed the segregation pattern and the difference of evolutionary changes. Guava varieties were separated into two main groups, one of them was made up of Chineese, Jelly, Kazi, Apple, L-49, Local-2 and Local-3. The other one was made up of Local-1, Poly, Kashi, Thai and Bombay. The highest genetic distance between Apple and Kazi peyara indicate that these varieties might be interesting in breeding programme for improving trait of interest. This scientific information could be used for further improvement of guava.

Keywords: Polymorphic, Genetic diversity, RAPD, Guava

Location: Germplasm Center of Khulna University and Khulna University campus, Bangladesh.

Start Date:

End Date:

Program Area: Variety and Species

Commodity: Guava

Objectives:

To assess the morphological variations through phenotypic study, to investigate the genetic variability at the genomic level and to select parents for better and more production.

Methodology:

Plant material: Twelve varieties of Guava were collected from Germplasm Center of Khulna University and Khulna University campus, Bangladesh. Genomic DNA extraction: The modified CTAB method (Doyel & Doyel, 1987) was adopted to extract genomic DNA from the plant leaves at Plant Biotechnology Lab of Khulna University, Khulna. In this experiment, 0.1g leaves were used instead of 0.5-1.0g leaves. Liquid nitrogen and sterile sand were avoided during grinding. For genomic DNA extraction isolation buffer, lysis buffer and CTAB buffer were used separately instead of CTAB isolation buffer. PCR reaction and RAPD analysis: Genomic DNA polymorphism was determined by the random amplified polymorphic DNA (RAPD) method (Williams et al., 1990). Five 10 mer primers and three 12 mer primers were used for DNA amplification (Table 2). Amplification was performed in sterile 0.2 ml Eppendorf tubes in 10 μl reaction volume. One microliter (2ng) of genomic DNA was mixed with 9 μl of PCR master mixture containing PCR buffer with additional 1.5 mM MgCl₂; 200 μM each of dATP, dCTP, dGTP, dTTP; 0.004 μM primer; 2 units of Taq DNA polymerase (Bioneer corporation, Korea). Amplification was conducted in a thermocycler (Eppendorp master cycler) for 5 min at 94°C (preliminary denaturation), the thermal cycle was 1 min at 94°C (denaturation), 1 min at 35°C (annealing) and 2 min at 72°C (elongation). This cycle was repeated for 30 times and an elongation period was carried out for 5 min at 72°C. Finally the reactions were held at 4°C. After PCR reaction, amplified products were analyzed by electrophoresis in 2% agarose gel. A Lambda/Hind III marker was run for each agarose gel, for estimation of fragment size. The gel was stained and visualized under UV Transilluminator. Gels were photographed using a gel documenter fitted with camera. Each accession was scored for the presence (1) or absence (0) of a particular amplification product. Data was analyzed by the software component 2.0 (CPW3) for phylogenic analysis. The unweighted pair group method with arithmetic averages (UPGMA) based dendrogram was constructed from distance matrix. The possible pair- wise genetic distance values were calculated according Nei & Li, (1979) d_xy = 1 – [2n_xy / (n_x-n_y)], where, n_x and n_y are the numbers of bands amplified in individuals x and y, respectively, and n_xy is the number of bands shared by those individuals. The distances between each pair of accessions were used to construct a dendrogram using the unweighted pair group method with arithmatic averages (UPGMA). Morphological features among the varieties were obtained by measuring the tree, leaves and fruits from randomly selected samples with assistance of the horticulture expert of Agro technology discipline, Khulna University. Fruit height was measured in meter. Ten mature fruits were collected for each variety. Fruit length and diameter were measured and the final value was obtained from average of those values. Fruits shapes were measured by adjusting the fruit shape chart.

Source of Information: Jahangirnagar University J. Biol. Sci. 7(2): 89-98, 2018 (December)

Article Link (Online Journal):

Funding Source:

Budget:

Total Budget (Tk.) :

Achievement/Findings:

From the above result Guava varieties were separated into two main groups, one of them was made up of Chineese, Jelly, Kazi, Apple, L-49, Local-2 and Local-3. The other one was made up of Local-1, Poly, Kashi, Thai and Bombay. The highest genetic distance between Apple and Kazi peyara indicate that these varieties might be interesting in breeding programme for improving trait of interest. This scientific information could be used for further improvement of guava.

Knowledge Management: Journal