Department of Biochemistry, University of Georgia, 30602, Athens, GA, USA.
Plant Mol Biol. 1983 Jul;2(4):199-206. doi: 10.1007/BF01578379.
The developmental time period and the magnitude of expression of the genes for the principal cottonseed storage proteins have been measured by several means. RNA was extracted from cotton cotyledons at stages during embryogenesis and the relative amounts of the mRNAS for these proteins were determined by cell-free translation in the wheat germ system and by dot and northern hybridization of the RNA with cloned cDNA probes representing the three subfamilies of the major storage protein genes. The rates of reassociation in solution of some of the RNAs with one of the cDNA clones were also determined. Data from all four procedures show that the storage protein mRNAs are demonstrable in very small embryo cotyledons, rapidly reach a high abundance level that is maintained during most of embryo growth, and then fall precipitously in amount in the last days of embryogenesis. The expression of all three gene subfamilies appears coordinate.Further, cDNA reverse transcribed from the poly(A)(+) mRNA from a stage of maximum storage protein synthesis was hybridized to saturation with cDNA clones representing each of the subfamilies. These data indicate that the mRNAs for two of the families reach the same relative level in the total mRNA population which is about 15% of the total mRNA mass. The mRNA of the third subfamily comprises only 5% of the total mRNA mass at this stage. This apparent 3∶3∶1 ratio of mRNAs does not change during the period of storage protein synthesis. Based on the amounts of the storage protein species in the mature seed, the mRNAs of each subfamily appear to be translated to the same extent during embryogenesis.
已经用多种方法测量了主要棉籽贮藏蛋白基因的发育时间和表达量。在胚胎发生过程的各个阶段从棉花子叶中提取 RNA,并通过无细胞翻译系统在小麦胚乳中测定这些蛋白质的 mRNAs 的相对含量,以及用代表主要贮藏蛋白基因三个亚家族的克隆 cDNA 探针进行 RNA 的斑点和 northern 杂交。还测定了一些 RNA 与一个 cDNA 克隆在溶液中的复性速率。所有四个程序的数据均表明,贮藏蛋白 mRNAs 在很小的胚胎子叶中即可检测到,迅速达到高水平,在胚胎生长的大部分时间内保持不变,然后在胚胎发生的最后几天急剧下降。所有三个基因亚家族的表达似乎都是协调的。此外,用来自最大贮藏蛋白合成阶段的 poly(A)(+)mRNA 反转录的 cDNA 与代表每个亚家族的 cDNA 克隆进行杂交,达到饱和。这些数据表明,两个家族的 mRNAs 在总 mRNA 群体中的相对水平相同,约占总 mRNA 质量的 15%。该亚家族的 mRNA 仅占该阶段总 mRNA 质量的 5%。在贮藏蛋白合成期间,这种明显的 3∶3∶1 mRNAs 比例没有改变。根据成熟种子中贮藏蛋白的含量,每个亚家族的 mRNAs 在胚胎发生期间的翻译程度似乎相同。