Systematic and benchmarking studies of pipelines for mammal WGBS data in the novel NGS platform.

BACKGROUND: Whole genome bisulfite sequencing (WGBS), possesses the aptitude to dissect methylation status at the nucleotide-level resolution of 5-methylcytosine (5-mC) on a genome-wide scale. It is a powerful technique for epigenome in various cell types, and tissues. As a recently established next-generation sequencing (NGS) platform, GenoLab M is a promising alternative platform. However, its comprehensive evaluation for WGBS has not been reported. We sequenced two bisulfite-converted mammal DNA in this research using our GenoLab M and NovaSeq 6000, respectively. Then, we systematically compared those data via four widely used WGBS tools (BSMAP, Bismark, BatMeth2, BS-Seeker2) and a new bisulfite-seq tool (BSBolt). We interrogated their computational time, genome depth and coverage, and evaluated their percentage of methylated Cs. RESULT: Here, benchmarking a combination of pre- and post-processing methods, we found that trimming improved the performance of mapping efficiency in eight datasets. The data from two platforms uncovered ~ 80% of CpG sites genome-wide in the human cell line. Those data sequenced by GenoLab M achieved a far lower proportion of duplicates (~ 5.5%). Among pipelines, BSMAP provided an intriguing representation of 5-mC distribution at CpG sites with 5-mC levels > ~ 78% in datasets from human cell lines, especially in the GenoLab M. BSMAP performed more advantages in running time, uniquely mapped reads percentages, genomic coverage, and quantitative accuracy. Finally, compared with the previous methylation pattern of human cell line and mouse tissue, we confirmed that the data from GenoLab M performed similar consistency and accuracy in methylation levels of CpG sites with that from NovaSeq 6000. CONCLUSION: Together we confirmed that GenoLab M was a qualified NGS platform for WGBS with high performance. Our results showed that BSMAP was the suitable pipeline that allowed for WGBS studies on the GenoLab M platform.

Isolation and characterization of OsMY1, a putative partner of OsRac5 from Oryza sativa L.

OsRac5 belongs to the rice Rho of plants family, and acts as the molecular switch in the signal pathway which is pivotally involved in the rice fertility control. One of its putative partners, OsMY1, was isolated by yeast two-hybrid screening from rice panicle cDNA library. Bioinformatics analysis shows that OsMY1 contains a coiled-coil domain which generally appeared in the partners of Rho GTPases. By yeast two-hybrid assay, it is confirmed that OsMY1 binds both the wild type (WT) and constitutively active (CA) OsRac5, but does not interact with dominantly negative OsRac5. In addition, the interactions between OsMY1 and WT-OsRac5 or CA-OsRac5 in vivo are demonstrated by bimolecular fluorescence complementation assay. Using PCR-mediated sequence deletion and point mutation of OsMY1, the interaction between OsMY1 and OsRac5 was identified to be mediated by the coiled-coil domain in OsMY1, and their binding was quantified by O-nitro-phenyl-ß-D-galactopyranoside assay. Real-time PCR shows that OsMY1 and OsRac5 are coordinately expressed in rice leaves and panicles with similar expression patterns. Our results suggest that OsMY1 is an important target of OsRac5 and that these two genes are involved in the same biological processes in rice growth and development.

Tillering and panicle branching genes in rice.

Rice (Oryza sativa L.) is one of the most important staple food crops in the world, and rice tillering and panicle branching are important traits determining grain yield. Since the gene MONOCULM 1 (MOC 1) was first characterized as a key regulator in controlling rice tillering and branching, great progress has been achieved in identifying important genes associated with grain yield, elucidating the genetic basis of yield-related traits. Some of these important genes were shown to be applicable for molecular breeding of high-yielding rice. This review focuses on recent advances, with emphasis on rice tillering and panicle branching genes, and their regulatory networks.