Molecular And Cytological Analyses Of Synaptonemal Complex Protein ZYP1 In Autopolyploid Arabidopsis Thaliana And Brassica Rapa

Maintenance of stable meiosis in polyploids is obligatory for controlled pairing and synapsis of more than two sets of homologous chromosomes, but such mechsnism remains under debate. Since the transverse filament protein ZYP1 is a critical component participating in synapsis of homologous chromosome and a timely prophase progression during meiosis. The present study investigated expression pattern of ZYP1 in autotetraploid Arabidopsis thaliana and Brassica rapa in contrast with diploid counterparts.It has provided some basic knowledge for study polyploidy regulation mechanism, by comparing the meiotic procress of diploid and autotetraploid. Immunofluorescence staining was performed by preparing the ZYP1 polyclonal antibody. The dynamic changes in the pairing and synapsis of homologous chromosome was traced by fluorescent labeling antibody ZYP1, and comparison of the fluorescence signal in diploid and autotetraploid Arabidopsis thaliana and Brassica rapa was made. Quantitative real-time PCR(qRT-PCR) was conducted to comparatively analyze the expression level of ZYP1 involved in autoteraploid Arabidopsis thaliana and Brassica rapa in comparison with diploid counterparts. The key synaptic element AtZYP1 protein labeled with fluorescent group is used to trace the dynamic progression of homologous chromosome pairing and synapsis during meiotic prophase by immunofluorescent staining, and a comparison of flourescence signal is made in Arabidopsis thaliana and Brassica rapa. Total protein in autoterploid and diploid Brassica rapa was extracted from the young inflorescences and separated on SDS-PAGE, then Western blot was made to make a comparison of the ZYP1 expression level in autotetraploid and diploid Brassica rapa. All above, we got some conclusions:1. The Arabidopsis thaliana ZYP1 prokaryotic expression vector was successfully constructed, and the optimized protein expression induction condition in E.coli was 1.0 mmol/l IPTG at 37 ℃. The polyclonal antibody had high specificity and sensitivity. It was able to detect and locate the antigen of ZYP1 protein in plant cells by immunofluorescence staining. This provide a guarantee for the later research.2. The dynamic changes of the fluorescence signal of ZYP1 in autoteraploid Arabidopsis thaliana and Brassica rapa were basically consistent with diploid counterparts. Chromosome surrounded together and chromosome centromere was more, due to the doubling, that repreasents the higher synapsis frequency in autotetraploid.3. ZYP1 flourescence signal was firstly observed during early-leptotene and gradually increased at zygotene, they tend to be more mature at pachytene. The flourescence signal then quickly dissipate from late-pachytene to diplotene and gradually disappeared in diakinesis. ZYP1 flourescence signal in autoteraploid Arabidopsis thaliana and Brassica rapa appeared more than in diploid counterparts. At the same time, the Real- time PCR analysis results demonstrated that the ZYP1 gene expression also significantly improved. The results of Western blot showed that the expression of ZYP1 protein in autoteraploid was higher.The results of Western blot showed that the expression of ZYP1 protein was higher than diploid counterparts. Therefore, the precise mechanism controlling pairing and synapsis of homologous chromosomes during meiosis likely evolved in polyploids via coordinate regulation of synaptic element ZYP1 protein to ensure genome fidelity after polyploidization.