Cytological Observation and Expression Analysis of Genes Involved in Anther Tetrad Releasing of a Thermo-sensitive Male Sterile Line SP2S in Brassica napus L.

doi:10.7668/hbnxb.2014.02.007

Abstract

Abstract: Thermo-sensitive genic male sterile line of Brassica napus plays an important role in two-line breeding. In this study, the new thermo-sensitive genic male sterile line of Brassica napus and its near isogenic line SP2F were used. The degradation of callose in SP2F and SP2S anthers was observed by aniline blue staining. The total RNA was extracted from different developmental stages of bud of lengths of 0-1,1-2,2-3,3-4 mm and reversely transcribed to cDNA. Amplification, sequencing and Real-time PCR analysis of genes involved callose dis- integration in microspore separation between SP2F and SP2S. The results indicated that: The callose of the SP2S was not degraded at tetrad stage, which made the microspores adhesion with each other and then pollen aborted. The cloned gene BnMSR66 belongs to plant β-1,3-glucanase genes and share a high similarity with other plants. The full length of BnMSR66 is 445 bp, and the ORF is 390 bp, encoding 130 amino acids, no difference between SP2S and SP2F. The expression of five genes, including BnMSR66 and BnA6 involved in callose dissolution, and three QRT genes (BnQRT1, BnQRT2, BnQRT3) control microspores separation after meiosis and produce mature pollen tet- rads, at four stages (PMC and meiosis stage, tetrad to the microspore released stage, uninucleate microspore, and the bicellular microspores stage, buds of 0-1, 1-2, 2-3, 3-4 mm length) were quantitatived by Real-time PCR, showed that all the five genes were significantly up-regulated in SP2S than in SP2F. Results of both cytological ob- servation and differential expression analysis exhibited that deformation and dysfunction of the tapetum lead to ab- normal expressing of the genes involved in microspore separation and microspore abortion.

Key words: Brassica napus L., Thermo-sensitive male sterility, Cytology, Callose, Real-time fluorescent quanti- tative PCR

CLC Number:

HU Yu-mei, XU Xian-feng, YU Cheng-yu, GUO Ying-fen, GE Juan. Cytological Observation and Expression Analysis of Genes Involved in Anther Tetrad Releasing of a Thermo-sensitive Male Sterile Line SP2S in Brassica napus L.[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2014, 29(2): 38-44. doi: 10.7668/hbnxb.2014.02.007.

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