臺灣博碩士論文加值系統

本試驗以台農67號品種的水稻為材料，構築10天胚之cDNA庫。並以10天胚
與成熟胚的mRNA合成單股cDNA為探針，差異性篩選出10天胚之cDNA庫中10
天胚特有的基因。其目的為 (1)選殖與分離出水稻胚10天獨特的基因，
(2)分析此基因在胚發育過程之表現特性， (3)探討此基因的結構經差異
性雜交篩選十二萬個溶菌斑，初步選出 828個具有十天胚差異表現的溶菌
斑。將初步篩選的溶菌斑殖系，轉換成質體，並重複進行南方轉漬差異性
雜交的篩選，最後獲得29個十天胚特有的殖系。所選殖的29個殖系，以其
cDNA片段做成 32-P 標誌的探針，分別與 10、15天、成熟胚和葉片之
mRNA做北方轉漬雜交，試驗結果可看出這29個殖系在10天胚之表現均較15
天、成熟胚和葉片組織內表現量高。北方轉漬雜交分析pRE4、pRE22、
pRE8、pRE20和pRE2等 5個選殖系，在胚體的時序及在組織織表現的特性
，顯示約在授粉後第 9天的胚體表現度較強，在 7天、10天、11天、15天
的胚表現稍弱，在成熟胚、葉片及胚乳組織的表現則相當不明顯。將篩選
得到的29個殖系，自3''端進行核酸定序，解讀 150-300個核酸序列，經比
對殖系核酸序列的相似性，將所選殖系分成四組，前三組的各個殖系之間
相似性極高，可能為同一基因或基因族；而第四組的各殖系間相似性低，
可能為不相同的基因。染色體組DNA 之南方墨點分析的結果，由呈現單一
或少數的雜交條帶訊號，顯示選殖系在染色體上是為單一或是少數的基因
座。另外，以選取的各組代表之殖系為探針，與台農67號、lomello 之品
種水稻、玉米、紅豆、小白菜、花椰菜、蕹菜和馬鈴薯等作物之三天幼苗
的RNA ，進行北方轉漬雜交，結果並未偵測到有雜交之訊號，顯示選殖系
在上述的不同的植物幼苗中，並無此基因的表現，故選殖系應是水稻胚發
育、胚特有的殖系。

The objectives of the current research are to isolate the
embryo- specific genes from 10-day embryos of rice, to examine
the spatial- specific and temporal-specific expression of
selected genes, and to study the genomic organization of these
genes. A cDNA library was constructed from 10-day embryos of
rice (Oryza sativa L. cv. Tainung 67). Eight-hundred and
twenty-eight plaques that hybridized with [32-P]-labeled single-
strand cDNA of 10-day embryos but not of mature embryos (45
days after pollination), were preliminary selected. Following
in vivo excision of selected phages and repeating DNA dot blot
hybridiation, 29 cDNA clones were further selected. The
results of northern slot hybridization indicated that the the
expression of 29 cDNA clones were higher in 10-day embryos than
in 15-day embryos, and there little expression in the mature
embryos or leaves of rice. The results of northern blot
hybridization showed that the expression of pRE4、 pRE22、 pRE
、 pRE20 and pRE2 were highest in 9-day embryos among 7-, 10-,
11, and 15-day embryos, and there were little expressions in
the mature embryos or leaves of rice. We were not able to
detect any expression of selected genes in the 3-day seedling
of rice, corn, azuki bean, pai-tsai, cauliflower, water
convolvulus and potato. Thus, the data indicated that selected
clones are 10-day embryos abundant and/or specific genes.
According to 3''end nucleotide sequencing, the selected 29 cDNA
clones could be divided into seven different genes or gene
families. The results of Southern slot blot hybridization of
rice genomic DNA indicated that the selected genes are the
single or a few copies in the chromosome.

The objectives of the current research are to isolate the
embryo- specific genes from 10-day embryos of rice, to examine
the spatial- specific and temporal-specific expression of
selected genes, and to study the genomic organization of these
genes. A cDNA library was constructed from 10-day embryos of
rice (Oryza sativa L. cv. Tainung 67). Eight-hundred and
twenty-eight plaques that hybridized with [32-P]-labeled single-
strand cDNA of 10-day embryos but not of mature embryos (45
days after pollination), were preliminary selected. Following
in vivo excision of selected phages and repeating DNA dot blot
hybridiation, 29 cDNA clones were further selected. The
results of northern slot hybridization indicated that the the
expression of 29 cDNA clones were higher in 10-day embryos than
in 15-day embryos, and there little expression in the mature
embryos or leaves of rice. The results of northern blot
hybridization showed that the expression of pRE4、 pRE22、 pRE
、 pRE20 and pRE2 were highest in 9-day embryos among 7-, 10-,
11, and 15-day embryos, and there were little expressions in
the mature embryos or leaves of rice. We were not able to
detect any expression of selected genes in the 3-day seedling
of rice, corn, azuki bean, pai-tsai, cauliflower, water
convolvulus and potato. Thus, the data indicated that selected
clones are 10-day embryos abundant and/or specific genes.
According to 3''end nucleotide sequencing, the selected 29 cDNA
clones could be divided into seven different genes or gene
families. The results of Southern slot blot hybridization of
rice genomic DNA indicated that the selected genes are the
single or a few copies in the chromosome.