RNA-Seq Uncovers SNPs and Alternative Splicing Events in Asian Lotus (Nelumbo nucifera).

Mei Yang,Pingfang Yang,Liming Xu,Yanling Liu,Peng Xu

doi:10.1371/journal.pone.0125702

Mei Yang, Pingfang Yang + Show 3 more

Open Access

https://doi.org/10.1371/journal.pone.0125702

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Journal: PloS one	Publication Date: Apr 30, 2015
Citations: 25	License type: CC BY 4.0

Affiliation: Wuhan Botanical Garden, Chinese Academy of Sciences

Abstract

RNA-Seq is an efficient way to comprehensively identify single nucleotide polymorphisms (SNPs) and alternative splicing (AS) events from the expressed genes. In this study, we conducted transcriptome sequencing of four Asian lotus (Nelumbo nucifera) cultivars using Illumina HiSeq2000 platform to identify SNPs and AS events in lotus. A total of 505 million pair-end RNA-Seq reads were generated from four cultivars, of which 86% were mapped to the lotus reference genome. Using the four sets of data together, a total of 357,689 putative SNPs were identified with an average density of one SNP per 2.2 kb. These SNPs were located in 1,253 scaffolds and 15,016 expressed genes. A/G and C/T were the two major types of SNPs in the Asian lotus transcriptome. In parallel, a total of 177,540 AS events were detected in the four cultivars and were distributed in 64% of the expressed genes of lotus. The predominant type of AS events was alternative 5’ first exon, which accounted for 41.2% of all the observed AS events, and exon skipping only accounted for 4.3% of all AS. Gene Ontology analysis was conducted to analyze the function of the genes containing SNPs and AS events. Validation of selected SNPs and AS events revealed that 74% of SNPs and 80% of AS events were reliable, which indicates that RNA-Seq is an efficient approach to uncover gene-associated SNPs and AS events. A large number of SNPs and AS events identified in our study will facilitate further genetic and functional genomics research in lotus.

Highlights

Lotus belongs to Nelumbonaceae, a small plant family with only one genus, Nelumbo, and two species: N. nucifera and N. lutea [1]
Single nucleotide polymorphism (SNP) markers could meet the needs on both marker density and genome coverage, and have been applied in linkage mapping and genome wide association studies (GWAS) in many species, for instance, Arabidopsis [5, 6], rice [7, 8], maize [9,10,11], soybean [12], sunflower [13, 14] and Cucurbita [15]
After filtering with NGS QC Toolkit [44], a total of 72.68, 74.85, 184.93, and 172.57 million high-quality reads in length of 100 bp were generated from ‘BG’, ‘WR1’, ‘ZO’, and ‘Red Lingxiao’ (RL)’, respectively

Summary

Introduction

Lotus belongs to Nelumbonaceae, a small plant family with only one genus, Nelumbo, and two species: N. nucifera (distributed in Asia, Australia, Russia) and N. lutea (distributed in eastern and southern North America) [1]. RNA-Seq on Illumina platform could generate redundant transcriptome sequences with high read depth and is a powerful way of identifying large scale SNPs from transcribed regions in the genomes [17,18,19,20]. The AS events have been identified from expressed sequence tags (ESTs) in lotus [37], the landscape of AS has not been explored from lotus RNA-Seq transcriptome data.

Results

Conclusion