Transcriptome analysis of two buffalograss cultivars

Abstract

Background: Buffalograss [Buchloë dactyloides (Nutt.) Engel. syn. Bouteloua dactyloides (Nutt.) Columbus] is a United States native turfgrass species that requires less irrigation, fungicides and pesticides compared to more commonly used turfgrass species. In areas where water is limited, interest in this grass species for lawns is increasing. While several buffalograss cultivars have been developed through buffalograss breeding, the timeframe for new cultivar development is long and is limited by a lack of useful genetic resources. Two high throughput next-generation sequencing techniques were used to increase the genomic resources available for buffalograss.

Results: Total RNA was extracted and purified from leaf samples of two buffalograss cultivars. '378' and 'Prestige' cDNA libraries were subjected to high throughput sequencing on the Illumina GA and Roche 454 Titanium FLX sequencing platforms. The 454 platform (3 samples) produced 1,300,885 reads and the Illumina platform (12 samples) generated approximately 332 million reads. The multiple k-mer technique for de novo assembly using Velvet and Oases was applied. A total of 121,288 contigs were assembled that were similar to previously reported Ensembl commelinid sequences. Original Illumina reads were also mapped to the high quality assembly to estimate expression levels of buffalograss transcripts. There were a total of 325 differentially expressed genes between the two buffalograss cultivars. A glycosyl transferase, serine threonine kinase, and nb-arc domain containing transcripts were among those differentially expressed between the two cultivars. These genes have been previously implicated in defense response pathways and may in part explain some of the performance differences between 'Prestige' and '378'.

Conclusions: To date, this is the first high throughput sequencing experiment conducted on buffalograss. In total, 121,288 high quality transcripts were assembled, significantly expanding the limited genetic resources available for buffalograss genetic studies. Additionally, 325 differentially expressed sequences were identified which may contribute to performance or morphological differences between 'Prestige' and '378' buffalograss cultivars.

Figure 1

Species distribution of the top BLAST hits for the ‘Prestige’ sequences. Transcripts were compared to the NCBI nr database (A), and also to model Liliopsida plants from Ensembl (B).

Figure 2

Histogram of Unigene clusters in ‘Prestige’ and ‘378’ cultivars. X-axis lists how many transcripts are in a unigene, and Y-axis lists how many unigenes are of that size.

Figure 3

The number of transcripts versus the coverage of blast alignments to subject protein sequences.

Figure 4

Scatter plot displaying percent length of ‘Prestige’ transcripts aligning to percent length of ‘378’ transcripts. Transcripts shown are those that were a reciprocal match and significant expression change was detected.

Figure 5

Number of significantly expressed genes in each of the Level 3 GO biological processes. Transcripts with higher expression in ‘Prestige’ than in ‘378’ are in red. Transcripts with higher expression in ‘378’ than in ‘Prestige’ are in green.