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mmunolocalization of Desiccation Associated LLA-32 Protein and Characterization of Pollen Germination Associated Genes in Lily Pollen
低溫儲存過的百合花粉萌發及生長速率會比新鮮花粉低，為了探討這個現象，在之前的研究中分別抽出儲存及新鮮花粉的全RNA，利用supression subtractive hybridization的方式，找出於低溫儲存過程中一些mRNA被降解的的基因，本論文中發現了45個基因片段；此外本實驗室也以上述方式發現63個基因片段，其中一個為單醣運輸蛋白質（LLPD22）。本論文以5'及3'-RACE的方式得到其cDNA全長共計1851 nt，並可轉譯出含505個氨基酸之蛋白質，分子量55.0 kDa，pI值為6.4。單醣運輸蛋白質在醣類運輸、轉換和能量產生的機制中有著重要地位。目前推測花粉管壁膜上之轉化酵素（invertase）可將蔗糖分解成果糖和葡萄糖，再經由單醣運輸蛋白質把果糖和葡萄糖由花粉管外送入花粉管內，以提供花粉管生長所需之能量及構築細胞結構之來源。|
LLA-32 is a stress-induced protein and is in association with desiccation in lily (Lilium longiflorum) pollen during the later stage of maturation. Previous study indicated that the accumulation of LLA-32 protein remained steady in the mature pollen, but it was gradually disappeared during pollen tube germination. To further investigate the possible function of LLA-32 during pollen maturation and pollen tube germination, immunolocalization with affinity-purified LLA-32 antisera was performed. Experimental results demonstrated that LLA-32 mainly localized in the pollen outer wall(exine), cytoplasm and nucleus of generative cell, and vegetative nucleus. Less labeling was found in vegetative cytoplasm and germination pollen tube, and was rare in the pollen inner wall(intine). The possible function of LLA-32 in mature lily pollen is protection of the pollen grain before pollen tube germination during desiccation. The germination and growth rates of lily pollen grain are dramatically reduced after low temperature storage. By using supression subtractive hybridizatio, the mRNAs degraded during -20℃ storage were isolated. In this studying, 45 cDNA clones were obtained. In the parallel experiment, one cDNA encoded for the monosaccharide transporter(LLPD22)obtained and chosen for further characterization. To obtain its full length, 5'- and 3'-RACE were perform. One full length of 1851 nucleotides was obtained and contained one opening reading with 505 a.a. Its estimated molecular weight and pI are 55 kDa and 6.4, respectively. It has been suggested that monosaccharide transporters play critical rules in sugars transport across plasma membrane. The invertase of pollen tube membranes may convert the sucrose into fructose and glucose, which are transported by monosaccharide transporters into pollen tubes to provide the energy and cell skeleton source during pollen tube growth.
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