Expression Analysis Of CACO Gene In Male Sterile Wheat Induced By Chemical Hybridizing Agent (CHA) SQ-1

Physiologically male sterility (PMS) induced by chemical hybrid agents (CHA) is an important and simple way in wheat hybrid production today. The utilization of CHA for hybrid yields has many merits, and the superior combinations between the conventional wheat varieties can be directly used for producing hybrid seeds. SQ-1 is a new type of wheat CHA which has wide spaying window and can lead to complete male sterility, and the side effects on the agronomic traits were not observed, it is also one of the best wheat CHAs all over the world. However, its mechanism for male sterility is still unclear, whether there exists difference in the levels of gene transcription and protein expression has become a focus in this field. In this study,'Xinong 1376'and'Xinong 2611'and their PMS lines were used as experimental materials to examine the activity of aconitase in the male anthers .Simultaneously, an EST with highly similarity to cytosolic aconitase from an AFLP-cDNA library was used as a querying probe to blast the Genbank databases, based on the assembled homologous cDNA sequence, the cytosolic aconitase (cACO) gene was amplified and characterized by PCR and sequence analysis. Furthermore, we have analyzed the expression characteristics of the gene cACO between male sterile and fertile anthers via semi-quantitative PCR in two different varieties to further discuss the expression profiles of the gene in the physiologically male sterile and fertile anthers .The main results obtained were as follows:1.In this research, the cACO activity was detected in control and SQ-1 treated wheat lines. The results showed that at late mononucleate stage the cACO activity in sterile anthers was extraordinary higher than that in fertile anthers, but at binucleate and trinucleate stages it was lower than that in fertile anthers. Therefore, the exceptional change of cACO activity may be changed the electron transport pathway of respiratory metabolism and cause the lack of energy supply and the shortage of intermediates needed in the binucleate and trinucleate stages during anther development. As a result, the altered activity of cACO may affect the anther development and eventually lead to male sterility, which is closely related to male sterility in wheat. 2.an EST with highly similar to cytosolic aconitase gene from the sterile AFLP-cDNA library was used as a querying probe to blast the Genbank databases, based on the assembled homologous, the cytosolic aconitase (cACO) gene was cloned and analyzed using semi-quantitative RT-PCR. The cloned cDNA sequence was 1368 bp in size and putatively encoded a peptide of 456 amino acids, and the assigned GenBank accession was GU475062.3. In the study, the expression pattern of cACO gene between male sterile and fertile anthers was analyzed via semi-quantitative RT—PCR in two different wheat varieties. The results showed the expression of cACO gene in sterile and fertile anthers were firstly upregulated and then decreased, but in contrast the expression level of cACO gene in sterile anthers was much higher than that in fertile anthers at the late mononucleate stage, and then it was much lower than that in fertile anthers at binucleate and trinucleate stages. It is suggested lacking of energy and necessary metabolic intermediates caused by abnormal expression of cACO gene at the key developmental stage eventually leaded to the abortion of pollens.