Cloning And Expression Analysis Of Skp1gene Related To Male Sterility In Wheat

The whea(tTriticum aestivum)is very impotant grain in world, also one of main material basis to our existing.It's an effective measure of increasing production and improving quality by heterosis. However, The theory of heterosis and research of practice with wheat are not deeper than maize, rice, rape and other crops.So far, study on the mechanism of male sterility is a significant breakthrough point for using heterosis.Summarized domestic and international wheat infertility type, It has two major aspects for studing male sterility including: male-sterility induced by Physiology and heredity. In the research, different scholars have different emphasis, they do these series study including the fertility of genes, difference of fertility proteins,physiological and biochemical metabolism. Ubiquitin proteasome pathway which can guarantee complex physilolgical and biochemical processes in proper operation is an important way to degradate redundancy protein in cells.One kind of low molecular weight protein SKP1(S-phase kinase-associated-protein 1)participating in ubiquitin proteasome pathway is found existing in eukaryotes broadly.It plays a decisive role in maintaining the biochemical function of SCF complex.The process of protein-degradating of ubiquitin mediated in cells is mainly regulated by SCF related with regulation in cell cycle.And at the same time, many physiological processes depends on SCF complex.In the study, male sterility intruduced by chemical hybrid agent SQ-1, the anther of xinong-2611 divided into two samples-the spraying and unspraying as the test object. From the former study,we find that male sterile induced by SQ-1 happens from the later of mononuclear to binuclear stage.So take cDNA of binuclear stage of the two samples as the test object,and then the primer of SKP2 gene is designed and amplificated based on sequence of SKP1 gene of arabidopsis from NCBI GeneBank.Next, make a homology comparison of the above sequence,and at the same time ,make an expression of quantification of SKP1 gene during mononuclear stage,binuclear stage and trinuclear stage,respectively in 2611.The study will lay the theoretical foundation for revealing the mechanism of male-sterile induced by physiology. The result is as follows via the experiment:(1)expression of SKP1 is related with physiological type male sterility by SQ-1 treated in wheat (2)used total RNA extracted from anthers in binuclear to strand cDNA by revearse transcrition,and then gene sequences(852bp) are finally gained. by comparison and judgement,the SKP1 gene sequence in binuclear stage of wheat2611 is found firstly and would to been submitted to NCBI GeneBank(3)Making a comparison of the two sequences achieved above with DNAman software,the result reveals that all bases remain unchanged except for the No.477 base.Through contrasting the protein sequences translated from the different bases with PrimerPrimer5.0 software,only one different base can't cause difference of protein.Conclusion: transcription of SKP1 gene remains uninfluenced on both test object-the spraying and unspraying, in other words,SQ-1 can't cause difference at transcriptional level on wheat.(4)Aalysis the expression of SKP1 gene of anthers in sterile plant and fertile plant induced by physiology respectively via fluorescence quantitatiye PCR technique. The results suggest that the expression of the gene all appears an decreasing tendency in three periods(mononuclear, binuclear and trinuclear)during the development of anthers in sterile and fertile plant. It's obvious that the expression of the gene in sterility plant during this three periods was dramatically inhibited compared with that in fertile plant. As a result,it can be easily supposed that influencing the ubiquitin-proteasome proteolytic pathway in the development of pollen grains,down-regulated SKP1 gene expression can lead to male sterility.