Characteristic Expression Of CDNAs Induced During Late Zygotene-pachytene Stages Of Microsporocytes In Lily Anthers

Prophaseâ… of plants is a complicated process,involving late zygotene and pathytene,which are the crucial stage to complete synapsis and recombination.The Lilium davidii var.willmottiae(E.H.Wilson) Raffill was chosen as research object, we constructed the EST libraries by suppression subtractive hybridization(SSH) of anther depending on the stages of pollen mother cells in late zygotene-pachytene,and some clones expression strengthened significantly during the stage were obtained. Partial ESTs were analysed by Real-Time PCR and in situ hybridization to investigate the expression characterization.In order to acquire some useful information about the process of the prophaseâ… of plants,main results as follows:1.Forward and reverse suppressive subtractive hybridization(SSH) libraries were constructed by cDNAs prepared from anthers depending on the stages of pollen mother cells(late zygotene-pachytene stages versus the stages of leptotene and early zygotene,and late zygotene-pachytene stages versus the stages from diplotene to tetraspore).2.Reverse Northern dot blot was carried out to further screen the SSH libraries. Positive clones were identified and 132 ESTs expression strengthened significantly were sequenced and assembled.The results indicated that there were 34 unique ESTs. The cDNA fragment sequences were compared with datas in GenBank by BLASTN and BLASTX.18 ESTs exhibiting significant homologies to the datas found in GenBank.34 unique ESTs were categorized by aligning them with MIPS principles and GO datas.They were classified into eight different groups:metabolism,protein modification and signal transduction et al.3.Real-Time PCR was performed to confirm the expression levels of 12 selected ESTs during late zygotene-pachytene stages of microsporocytes in lily anthers and other tissues.4.The clone bh1-1-f9(LIM12 mRNA) was investigated by in situ hybridization. Combined the datas of Real-Time PCR,the results initially revealed the expression pattern of LIM12 during the stages of pollen mother cells in late zygotene-pachytene in lily anther.In this study,SSH were used for the screening of ESTs expression strengthened during the stages of pollen mother cells in late zygotene-pachytene in lily anther.In particular,a number of unknown genes in the library will be selected for future study to provide some useful information.