An allopolyploid formation consists of the two processes of hybridisation and chromosome doubling. Hybridisation makes a different genome combined in the same cell, and genome “shock” and instability occur during this process, whereas chromosome doubling results in doubling and reconstructing the genome dosage. Recent studies have demonstrated that small RNAs, mainly siRNAs and miRNAs, play an important role in maintaining the genome reconstruction and stability. However, to date, little is known regarding the role of small RNAs during the process of wide hybridisation and chromosome doubling, which is essential to elucidate the mechanism of polyploidisation. Therefore, the genetic and DNA methylation alterations and changes in the siRNA and miRNA were assessed during the formation of an allodiploid (genome: AB) and its allotetraploid (genome: AABB) between Brassica rapa (♀) and Brassica nigra (♂) in the present study.The phenotypic analysis exhibited that the allotetraploid had high heterosis compared with their parents and the allodiploid. The methylation-sensitive amplification polymorphism (MSAP) analysis indicated that the proportion of changes in the methylation pattern of the allodiploid was significantly higher than that found in the allotetraploid, while the DNA methylation ratio was higher in the parents than the allodiploid and allotetraploid. The high-throughput sequencing results obtained for the small RNAs showed that the expression levels of miRNAs increased in the allodiploid and allotetraploid compared with the parents, and the expression levels of siRNAs increased and decreased compared with the parents B. rapa and B. nigra, respectively. Moreover, the percentages of miRNAs increased with an increase in the polyploidy levels, but the percentages of siRNAs and DNA methylation alterations decreased with an increase in the polyploidy levels. Furthermore, 320 known and 52 novel miRNAs were obtained from the parents in both the allodiploid and allotetraploid. However, quantitative real-time polymerase chain reaction (qRT PCR) analysis showed that the expression levels of the targets genes were negatively corrected with the expressed miRNAs.The study showed that siRNAs and DNA methylation play an important role in maintaining the genome stability in the formation of an allotetraploid. The miRNAs regulate gene expression and induce the phenotype variation, which may play an important role in the occurrence of heterosis in the allotetraploid. The findings of this study may provide new information for elucidating that the allotetraploids have a growth advantage over the parents and the allodiploids.
Overall design: High throughput sequence of the parents (Brassica rapa and Brassica nigra) and their hybrids (allodiploid and allotetraploid)
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