Comparative transcriptome analysis reveals key genes associated with meiotic stability and high seed setting rate in tetraploid rice

Abstract

Background: Polyploid rice has a high yield potential and excellent nutritional quality. The development of polyploid rice remained critically limited for several decades due to low seed setting rate until the successful breeding of polyploid meiosis stability (PMeS) lines. To determine the mechanism responsible for meiotic stability and high seed setting rate of PMeS line, agronomic traits, pollen fertility and viability, and meiotic behaviors of PMeS and non-PMeS lines were investigated. Further, comparative transcriptome analysis was performed to identify genes associated with meiotic stability and high seed setting rate in PMeS line.

Results: The seed setting rate, fertile and viable pollen ratios of PMeS line were significantly higher than those of non-PMeS line. The PMeS line exhibited stable meiosis, and chromosomes mainly paired as bivalents, rarely as univalents and multivalents in prophase I. Few lagging chromosomes were observed in anaphase I. By contrast, the homologous chromosomes pairing was disorganized in the non-PMeS line, with low frequencies of bivalents and high frequencies of univalents and multivalents in prophase I, while more cells with increased lagging chromosomes were detected in anaphase I. Many differentially expressed genes (DEGs) between PMeS and non-PMeS lines were identified through comparative transcriptome analysis. Some meiosis-related genes were specifically investigated from all DEGs. Further, several meiotic genes were identified as candidate genes.

Conclusions: The study not only demonstrates the morphological, cytological, and molecular differences between the PMeS and non-PMeS lines, but also provides several key genes associated with meiotic stability and high seed setting rate in tetraploid rice.

Keywords: High seed setting rate; Meiotic genes; Meiotic stability; Tetraploid rice; Transcriptome analysis.

Fig. 1

Morphological characteristics of HN2026-4x and 9311-4x. A Plants; B Panicles; C Grains. Left: HN2026-4x; Right: 9311-4x; Bars: A = 20 cm; B = 5 cm; C = 1 cm

Fig. 2

The meiosis behavior of pollen mother cells in HN2026-4x and 9311-4x. A, B, C, D HN2026-4x; E, F, G, H 9311-4x. A, E Prophase I (Diakinesis); B, F Metaphase I; C, G Anaphase I; D, H Telophase I. The lagging chromosomes are indicated by arrows. Bar = 10 μm

Fig. 3

Analysis of pollen fertility and viability of HN2026-4x and 9311-4x. A Pollens of HN2026-4x stained by I₂-KI; B Pollens of 9311-4x stained by I₂-KI; C Pollens of HN2026-4x stained by FDA; D Pollens of 9311-4x stained by FDA; E Comparison of pollen fertility and viability; H: HN2026-4x; Y: 9311-4x; Significant differences were determined using independent samples t-test; *** and **** indicate significant differences at 0.001 and 0.0001 probability levels, respectively. Bar = 200 μm

Fig. 4

Volcano map of differentially expressed genes between HN2026-4x and 9311-4x. Each dot represents a gene, with turquoise dots representing differentially expressed genes and red dots representing non differentially expressed genes

Fig. 5

Gene ontology (GO) analysis of differentially expressed genes between HN2026-4x and 9311-4x. The percentage and number of genes in each subcategory are shown on the left and right y-axes, respectively. The GO subcategories are shown on the x-axis

Fig. 6

Clusters of orthologous groups (COG) classification of differentially expressed genes between HN2026-4x and 9311-4x. A total of 329 unigenes are classified into 22 COG categories

Fig. 7

Validation of six differentially expressed genes by qRT-PCR. H: HN2026-4x; Y: 9311-4x. Significant differences were determined using independent samples t-test; *, ***, and **** indicate significant differences at 0.05, 0.001, and 0.0001 probability levels, respectively