Cloning a somatic embryogenesis like-kinase (serk) gene of Paspalum notatum

PODIO, MARICEL; FELITTI S A; SIENA, L A; ORTIZ J P A

Buenos Aires - Argentina

Congreso; The 6th International Symposium on the Molecular Breeding of Forage and Turf; 2010

CLONING A SOMATIC EMBRYOGENESIS LIKE-KINASE (SERK) gene of Paspalum notatum Podio M.1,2, Felitti S.A.1, Siena L.A.1 y Ortiz J.P.A.1,2 1Laboratorio de Biología Molecular, Facultad de Ciencias Agrarias, UNR. Campo Experimental J. Villarino C.C. N° 14 (S2125ZAA) Zavalla- Santa Fe- Argentina. 2Instituto de Botánica del Nordeste (IBONE)- CONICET, Facultad de Ciencias Agrarias, UNNE. CC209 (3400) Corrientes, Argentina. maricelpodio@yahoo.com.ar Paspalum notatum is a rhizomatous species widely distributed in native grasslands from central Mexico to Argentina. Different ploidy levels were reported for this species. Sexual diploids are cross-pollinated due to self-incompatibility, while the tetraploid races reproduce by obligate aposporous apomixis. This type of reproduction involves the formation of unreduced megagametophytes from nucellar cells, and the development of the embryo by parthenogenesis. Endosperm formation requires the fertilization of the two polar nuclei (pseudogamy). Apomixis results in progenies that are genetically exact copies of the female plant. The SOMATIC EMBRYOGENESIS RECEPTOR-LIKE KINASE (serk) gene plays a fundamental role in somatic embryogenesis of several angiosperms and was associated with apomixis in Poa Pratensis.􀀁 It was proposed that Ppserk activation in nucellar cells of apomictic genotypes of P. pratensis is the switch that channels embryo sac development and that it could redirect signaling gene products to compartments other than their typical ones (somatic cells instead of a megaspore mother cell). The objective of this work was to isolate and characterize the gene serk from tetraploid P. notatum. Oligonucleotides were designed based on conserved regions of serk genomic sequences of P. pratensis, rice and maize. A fragment of 200 bp was amplified using these oligonucleotides on genomic DNA from genotypes Q4188 (sexual) and Q4117 (apomictic). A chromosome walking strategy was used for extending Pnserk from six noncloned genomic libraries. A fragment of 711 bp, containing homologous sequences to the exons 9 and 10 of Ppserk and the intronic flanking sequences, was isolated. Southern-blot hybridization indicated the presence of at least five copies of Pnserk in tetraploid genotype Q4117. Further extension experiments will allow the isolation of the complete genomic and cDNA sequences of Pnserk in order to functionally characterize this gene in the species.