To identify genes that play critical roles during male gametogenesis in Arabidopsis, we have isolated several pollen morphological mutants from a mutagenized seed pool generated with a T-DNA activation vector. In this study, we have focused on a mutant plant producing ~50% abnormal pollen grains including high levels of collapsed pollen at maturity. The pollen developmental analysis showed that the mutant pollen phenotype was first observed at tricellular stage. Interestingly, the mutation was only maintained as a heterozygote due to the severely reduced genetic transmission through both sexes. TAIL PCR analysis led to the identification of the responsible gene which encodes a conserved oligomeric golgi complex component-related protein (COGCC). RT-PCR analysis showed predominant expression of the gene in reproductive organs including developing spores. The gene identity was confirmed by the result that mutant plants harboring a T-DNA containing corresponding wild type gene produced less level of mutant pollen grains. Furthermore, confocal laser scanning microscopy using mature pollen expressing COGCC-RFP driven under the native promoter showed small punctate signals, which are likely to be from the Golgi complex. Further experiments for co-localization of the COGCC-RFP with the Golgi markers are underway.

Based on the results of microarray analysis we selected ten candidate genes that express in pollen at the early pollen developmental stage. By PCR amplification, the promoter region of these genes were amplified from rice genomic DNA (Nipponbare) and cloned into the destination pKGWFS7 vector via an entry vector, pDONR201. The characteristic of promoters were evaluated in Arabidopsis thaliana (Col-0) through GUS expression analysis. Fifty T2 plants respectively from each promoter were tested. Whole inflorescence of individual plant was stained with 1mM X-Gluc solution to observe tissue-specific GUS expression patterns. The results showed that all 10 promoters activated in pollen tissues. Among them six promoters expressed at the early developmental stage (unicellular) of pollen and the others expressed at both early (unicellular) and late pollen developmental stage (mature pollen). The results indicated that these promoters would be potential applicable for the studies of pollen function. Currently, we are performing these promoters analysis in rice transgenic plants as well as molecular characterization.

A diverse number of genes are involved in the floral transition and development to ensure the proper timing on the switch from vegetative to reproductive development in Arabiodopsis. MADS-box genes play a major role in floral development especially in the case of vernalization process, In this study we mapped a mutation in MAF5 encoding a MADS-domain protein which was reported to be up-regulated during vernalization and regulates flowering time. The mutant in MAF5 showed several pleiotropic phenotypes that includes semi-dwarfism, delayed senescence and abnormal pollen phenotype, High percentages of vacuolated and aborted pollen phenotype were observed in the mutant plant. Transmission efficiency showed that mutation from this gene was defective in both male and female gametes. Furthermore, gene expression analysis revealed that this gene was predominantly expressed in reproductive organs and gave a strong expression in the mature pollen which coincides with the defect in pollen phenotype. The results from this study provide some evidences on the additional role of MAF5 in pollen development however more specific approaches should be done to determine the specific stages of pollen development altered in this mutant.

국내 재배 유채의 육종을 위해서는 소포자배양을 통한 배발생법의 확립이 절실하나 대부분의 재배 품종이 배발생율이 낮고 품종별로 화뢰의 크기에 따른 소포자의 발달 상태가 다르기 때문에 이에 대한 조사가 반드시 필요하다. 따라서 본 연구에서는 국내 육성 유채 품종의 육종을 위한 기초 자료로서 품종 간 화뢰의 크기에 따른 생체 내 화분(핵)발육 상태를 비교, 관찰하였다. 1. 소포자 배양에 있어서 배 발생이 가능한 소포자 분리 시기는 1핵기말기 또는 2핵기초기가 효과적이며, 소포자 배양에 최적인 화뢰의 크기는 품종에 따라 차이가 있지만 3.5-4.0 mm이다. 2. 예외적으로 '탐라유채'는 3.5-4.0 mm의 크기에서 다수의 1핵기말기의 소포자와 2핵기초기의 소포자가 관찰되었으며, 화뢰크기가 더 큰 4.0-4.5 mm 크기에서도 2핵기초기의 핵이 관찰되어 화분발육상태가 늦게 진행되는 것을 알 수 있었다. 3. '목포64호'의 경우 3.0-3.5 m크기에서 1핵기초기와 1핵기중기 및 1핵기 말기의 소포자들이 관찰되었으나 다른 품종과 달리 2핵기 화분이 전혀 관찰이 되지 않았다. 'MS-B Line'의 경우는 2.5-3.0 mm 크기에서도 4분자포자가 관찰되어 초기 발육상태가 조금 늦게 일어나는 것을 볼 수 있었다.

Thirty-nine out of eighty-five barley varieties/strains survived until heading stage in the saline experimental field (0.03-0.05~%~;salt) and they were used for pollen study. Light and scanning electron microscopic observations revealed two distinctive types of barley pollens: one transparent and small in size and the other dark and larger. In addition, both types of pollens were stained with Alexander's stain and it was found that the smaller and transparent pollen was cytoplasm-devoid (CD) while the larger pollen was cytoplasm-rich (CR). Sixteen out of 39 barley varieties/lines grown in the saline soil had CR pollens, which were rarely observed in the barley plants grown in the non-saline soil. Moreover, it was observed that salt stress severely reduced seed setting in the varieties having degenerated pollens. These results suggest that salt stress affects the fertility of barley pollen. The sterile pollen was undersized and lack of cytoplasm probably due to abortion. Furthermore, a varietal difference existed in the response of pollen development to salt stress.