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Morphological and Molecular Characterization of Grasspea

Md. Rezwan Molla
Scientific Officer
Plant Genetic Resources Centre, Bangladesh Agricultural Research Institute(BARI), Joydebpur, Gazipur-1701, Bangladesh

M. A. N. Nazim-ud-Dowla
Scientific Officer
Plant Genetic Resources Centre, Bangladesh Agricultural Research Institute(BARI), Joydebpur, Gazipur-1701, Bangladesh

Md. Shefatur Rahman
Scientific Officer
Plant Genetic Resources Centre, Bangladesh Agricultural Research Institute(BARI), Joydebpur, Gazipur-1701, Bangladesh

Sharmin Rahman
MS student
Bangabandhu Sheikh Mujibur Rahman Agricultural University, Salna, Gazipur

Abstract/ Executive Summary:

Genetic diversity was conducted in 25 grasspea accessions for thirteen morphological characters and at molecular level using Randomly Amplified Polymorphic DNA (RAPD) marker. The analysis of variance for all the characters revealed highly significant variation among the accessions of grasspea except for days to 50% fruiting. A total of 84.58% variation was observed from PCA against first thirteen eigen values while only first two values were responsible for 53.25% variation. The inter cluster distance was maximum (17.038) between the clusters of V and III followed by the distance (12.861) between the clusters of IV and III and the lowest (2.6) distance between IV and I in morphological analysis. Molecular analysis also showed higher genetic distance among parents of these clusters like BD 3618 vs BD 3686 (0.8232), BD 3685 vs BD 3621(0.7178) and BD 3686-BD 3621(0.7178). In both analysis the higher inter genotypic distance was observed between BD 3621-BD 3686 as well as BD 3618-BD 3686 while the lowest distance was observed between BD 3686-BD 3638. BD 3621-BD 3686 and BD 3618-BD 3686 have separate cluster and showed higher distance in both morphological (1.9654) and (1.8179) and RAPD marker based Nei’s genetic distance (0.7178) and (0.8232) respectively.

Keywords: Lathyrus., morphological characters, RAPD marker, genetic distance

Location: Bangabandhu Sheikh Mujibur Rahman Agricultural University, Salna, Gazipur and Molecular Biology Laboratory, Plant Genetic Resource Centre, Bangladesh Agricultural Research Institute (BARI), Gazipur-1701

Start Date: 01-10-2009

End Date: 01-03-2010

Program Area: Variety and Species

Commodity: Plant

Objectives:

• To determine the diversity among different accessions at morphological and molecular level • To reveal variation in morphological traits and DNA level within each of the grasspea accessions

Methodology:

Morphological Characterization The experiment was conducted using 25 grasspea accessions collected from the PGRC, BARI. The experiment was laid out in a Randomized Complete Block Design (RCBD) with three replications. Plant to plant and line to line distances were 4cm and 45cm, respectively. Morphological data was recorded according to Descriptors for Lathyrus spp. (IPGRI, 2000). Molecular Characterization Young and actively growing fresh leaf tissues of 15 days old seedling were collected from 25 individuals (five individuals per accession and mix) for the isolation of genomic DNA. Total genomic DNA was isolated from grasspea leaves following SDS extraction, Phenol: Chloroform: Isoamyl alcohol purification and ethanol precipitation method. A set of four primers (OPB-04, OPC-05, OPC-09, OPD-02) were evaluated based on intensity of bands, consistency within individual, presence of smearing, and potential for genotype discrimination. DNA amplification was performed in an oil-free thermal cycler (Thermal cycler, Techne, TC 512) with initial mix was preheated at 94°C for 3 minutes followed by 45 cycles of 1 min denaturation at 94°C, 1 min annealing at 35°C and extension at 72°C for 2 minutes. After the last cycle, a final step of 7 minutes at 72°C was added to allow complete extension of all amplified fragments. After completion of cycling program, reactions were held at 4°C. PCR products from each sample were confirmed by running 1.4% agarose gel. The sizes of amplification products were estimated with 100bp DNA ladder. DNA bands were visualized by staining with ethidium bromide and photographed on a UV transilluminator using gel documentation system. Estimating polymorphic loci, Nei’s (1973) gene diversity, genetic distance (D) and constructing a UPGMA (Unweighted Pair Group Method of Arithmetic Means) dendrogram among the populations were constructed using computer program POPGENE (Version 1.31) (Yeh et al., 1999).

Source of Information: ANNUAL RESEARCH REPORT OF PLANT GENETIC RESOURCES CENTRE, BARI -2009-2010

Article Link (Online Journal):

Funding Source:

1. Government

Budget: Tk. 1,00,000.00

Total Budget (Tk.) : Tk. 1,00,000.00

Achievement/Findings:

Plant height, pods/plant and seed yield/ plant had the highest contribution towards the divergence among 25 accessions of grasspea. BD 3587, BD 3618, BD 3621, and BD 3622 were identified as good accession due to their desirable agronomic traits. Based on analysis the accessions were grouped into five clusters. From, morphological study, the inter cluster distance was maximum (17.038) between the cluster V and III followed by the distance (12.861) between cluster IV and III. Molecular analysis also showed higher genetic distance among some parents of these clusters like BD 3618-BD 3686 (0.8232), BD 3685-BD 3621(0.7178) and BD 3686-BD 3621(0.7178).The similar result from both of the analysis confirmed that morphological and molecular analysis were effective and repeatable.

Knowledge Management: Report/Proceedings