Identification of a SCAR marker linked to apomixis in buffelgrass (Cenchrus ciliaris L.)

Apomixis is a mode of asexual reproduction through seeds. The apomictic process bypasses both meiosis and egg cell fertilization, producing offspring that are exact genetic replicas of the mother plant. Cenchrus ciliaris (buffelgrass) is a valuable forage grass, which reproduces through aposporous apomeiosis. We identified one sexual plant in this obligate apomictic population. The objective of this work was to identify molecular markers linked to apospory trait through bulked segregant analysis. Facultative apomictic F1 plants obtained from open pollinated sexual plant were selfed to derive F2 population. Individual F2 plants were classified through embryo sac analysis as sexual, facultative apomictic or obligate apomictic. Two bulks namely, obligate apomictic and sexual were prepared by pooling DNA of individual identified plants and used in PCR analysis. Out of 208 primers tested, 20 were found to be polymorphic. A 600 bp RAPD amplicon specific to apomictic bulk, obtained with OPF-08 primer was cloned and sequenced. Based on the sequence information, a pair of SCAR (sequence characterized amplified region) primers was designed which generated a 600 bp amplicon (named OPF08-600) only in the apomictic plants. Southern hybridization of genomic DNA of apomictic and sexual plants using the OPF08-600 fragment as probe indicated the presence of two copies of the SCAR sequence in only the apomictic plants. Gel elution of restricted DNA fragments and PCR analysis confirmed the presence of duplicated copies of the SCAR sequence in the apomictic plants.