Genetic identification of kikuyu grass (Pennisetum clandestinum) cultivars by RAPD and ISSR techniques.

Abstract

Kikuyu grass (Pennisetum clandestinum), originally from tropical and subtropical African countries, is one of the most important pasture grasses in the higher rainfall regions of South Africa (of utmost importance in agriculture, to the economy and to the industry). However, different cultivars are poorly characterised on a morphological basis and diagnostic genetic markers are missing, which could substantially support successful breeding and could be of crucial importance for rural dairy farmers who largely depend on low cost pasture crops. This study is aimed at finding genetic markers for cultivars of kikuyu grass based on two different PCR based methods. Random Amplified Polymorphic DNA (RAPD) and Inter-Simple Sequence Repeat (ISSR) marker systems were employed to detect polymorphism and identify genetic relationships among forty (40) kikuyu lines from Cedara, Department of Agriculture near Hilton, KwaZulu-Natal. Thirteen (13) RAPD primers amplified a total of 144 reproducible bands of which 80 were polymorphic and fourteen (14) ISSR primers amplified a total of 90 markers of which 56 were polymorphic. The percentage of polymorphic bands detected by ISSR and RAPD was fairly similar (62.22 and 55.56 %, respectively). Band scoring was analysed and FastTree dendrograms were constructed using the raxmlGUI1.3 and viewed using FigTree v1.4.0 analysis programs. Cluster analysis of the thirteen (13) informative RAPD primers produced an unrooted tree which grouped the forty cultivars into 8 distinct clusters (comprised of 3 larger clusters and 5 smaller ones) and 9 independent branches. The cluster analysis of the fourteen (14) informative ISSR primers produced an unrooted tree which grouped the forty cultivars into 4 distinct clusters (made of 2 larger clusters and 2 smaller ones) and 6 independent branches. Some cultivars were elucidated to share common clusters in dendrograms of both techniques and authenticate their genetic relationship among other cultivars. Cultivars sharing the same cluster (cluster 1) in both, the ISSR dendrogram and the RAPD dendrogram were cultivar 30, cultivar 40, cultivar 35, cultivar 39, cultivar 38, cultivar 34, and cultivar 36. Moreover, cultivar 11, cultivar 20, cultivar 19, cultivar 16, cultivar 14, cultivar 13, cultivar 15, and cultivar 12 showed the same grouping in both dendrograms (cluster 6) and (cluster 4), with cultivar 13 and 14 sharing more similarity than any other cultivar within the cluster. The cluster analysis results for both molecular techniques showed that Kikuyu grass cultivars tested here harbour considerable genetic variation, as was expected from the results of other preliminary research work focussing on physiological characters.