Isolation and characterization of microsatellite markers for Brachiaria brizantha (Hochst. ex A. Rich.) Stap

Abstract

The first set of nuclear simple sequence repeat (SSR) loci for Brachiaria brizantha (Hochst. ex A. Rich.) Stap is described. A microsatellite-enriched library was constructed and 19 loci were characterized. About 13 SSR loci were found to be polymorphic and across-taxa amplification tests showed that six of them can be transferred to four other species of Brachiaria. This new SSR resource will be a powerful tool for population genetic studies of B. brizantha, for interspecific genetic studies within the genus Brachiaria, for mapping and for marker assisted selection in breeding. Keywords Brachiaria brizantha Á Microsatellite markers Á Genetic diversity Á TransferabilityBrachiaria brizantha (Hochst. ex A. Rich.) Stapf is a grass native to tropical Africa, which reproduces predominantly by apomixis and presents high variation in ploidy levels (2, 4, 5, 6 and 9X). Taxonomy within this genus is controversial. Trinius (1834) described Brachiaria as a subdivision of Panicum. Grisebach (1853) elevated Trinius's subdivision to the category of genus. Webster (1987) has transferred B. brizantha, as well as several other species traditionally included in Brachiaria to the related genus Urochloa. Irrelevant to the taxonomic allocation, Renvoize et al. (1996) demonstrated that this group of species has a highly diverse nature and highlighted its enigmatic status in terms of both the component species and relationships with other genera. Brachiaria brizantha is closely related to B. decumbens and B. ruziziensis, and specimens are at times difficult to distinguish.This first set of microsatellites for B. brizantha is useful not only for conservation of naturally occurring specimens in African savannas, but also for understanding genetic relationships between B. brizantha and its related species, and for further mapping and marker assisted selection in breeding.Genomic DNAs were extracted from freeze-dried leaf samples using the CTAB method (Doyle and Doyle 1987).

Development and validation of microsatellite markers for Brachiaria ruziziensisobtained by partial genome assembly of Illumina single-end reads

“…In B. brizantha [15], 96 clones from an enriched library were sequenced, 19 primer pairs were designed and tested, and 13 of those were polymorphic. A new set of 15 polymorphic primers for this species was published by Vigna et al [16], using the same enriched library.…”

Genetic Diversity and Population Structure of the Brachiaria brizantha Germplasm

“…The 15 new SSRs presented in this study, along with five previously reported loci (Jungmann et al 2009b), were used to characterize the genetic diversity of the 172 accessions and six commercial cultivars of B. brizantha preserved in the EBC germplasm collection ( Table 2). The degree of polymorphism found in these loci among 178 genotypes was highly correlated with the degree of PIC values obtained from 23 accessions.…”

“…This type of marker was also used by Ambiel et al (2008) to analyze the same materials studied by Machado Neto et al (2002). The most recent study used microsatellites to evaluate 23 accessions of this species together with one genotype of each of the following Brachiaria species: B. ruziziensis, B. humidicola, B. decumbens and B. dictyoneura (Jungmann et al 2009a). Despite B. brizantha being studied together with other Brachiaria species in these four works, an evaluation of a definitive number of accessions has not been conducted until now.…”

“…To date, a panel of 13 SSR markers has been developed by our research group for B. brizantha (Jungmann et al 2009b). Here, we present 15 new polymorphic microsatellites for this species and their application, along with five previously published markers, in analyzing the intraspecific genetic diversity and population structure in 172 accessions and six cultivars of this species that are conserved in the germplasm collection held at EBC.…”

Development of microsatellite markers for Brachiaria humidicola (Rendle) Schweick

“…A genomic library was constructed using one B. humidicola genotype, following the methodology of Billotte et al (1999) and according to Jungmann et al (2009). A total of 384 clones were sequenced in an ABI 377 automated sequencer (PE Applied Biosystems) using the BigDye Terminator Cycle Sequencing Kit (Applied Biosystems).…”