Department of Crop Genetics and Breeding, Institute of Crop Sciences, Chinese Academy of Agricultural Science, Beijing, China.
Beijing Key Lab of Plant Resource Research and Development, Beijing Technology and Business University, Beijing, China.
PLoS One. 2014 Jan 9;9(1):e85159. doi: 10.1371/journal.pone.0085159. eCollection 2014.
The recessive opaque-2 mutant gene (o2) reduces α-zeins accumulation in maize endosperm, changes the amino acid composition of maize kernels, induces an opaque endosperm, and increases the lysine content of kernels. The quality protein maize (QPM) inbred line CA339 (o2o2) and an elite normal inbred line liao2345 (O2O2) were used to construct o2 near-isogenic lines (NILs) by marker-assisted selection (MAS) using the co-dominant SSR marker phi057. Two specific o2 NILs were constructed, named liao2345/o2-1 and liao2345/o2-2. However, the kernel phenotypes of the two o2 NILs were different from each other. liao2345/o2-1 had the wild-type vitreous endosperm, which is similar to its recurrent parent liao2345, while the endosperm of liao2345/o2-2 was opaque, identical to typical o2 mutant individuals. In comparison to their recurrent parent liao2345, the lysine concentration of liao2345/o2-1 was similar and the lysine concentration in liao2345/o2-2 was doubled. SDS-PAGE analysis indicated that liao2345/o2-1 had the same zeins ratio as liao2345, whereas the zeins concentration of liao2345/o2-2 was markedly lower. Sequence and transcript abundance analyses indicated that the CDS of two o2 NILs are derived from CA339, but they have different promoters. The O2 transcript of liao2345/o2-2 is largely inhibited because of an rbg transposable element inserted between the TATA box and initiator codon of liao2345/o2-2. We concluded that different crossing-over patterns during the process of o2 NIL construction resulted in the different kernel phenotypes of the two o2 NILs. We surmise that the reversion of liao2345/o2-1 to wild type was due to the recombination with the wild type liao2345 promoter during introgression and backcrossing. The o2 mutant gene of donor (CA339) is a null mutant because of low O2 expression. However, its CDS probably encodes a protein with normal function which can maintain the normal accumulation of zeins in maize endosperm.
隐性 opaque-2 突变体基因（o2）减少了玉米胚乳中α-醇溶蛋白的积累，改变了玉米籽粒的氨基酸组成，诱导了不透明的胚乳，并增加了籽粒中的赖氨酸含量。优质蛋白玉米（QPM）自交系 CA339（o2o2）和优良的普通自交系 liao2345（O2O2）被用于通过共显性 SSR 标记 phi057 进行标记辅助选择（MAS）来构建 o2 近等基因系（NIL）。构建了两个特定的 o2 NIL，分别命名为 liao2345/o2-1 和 liao2345/o2-2。然而，这两个 o2 NIL 的籽粒表型彼此不同。liao2345/o2-1 具有类似于其轮回亲本 liao2345 的野生型玻璃态胚乳，而 liao2345/o2-2 的胚乳是不透明的，与典型的 o2 突变体个体相同。与它们的轮回亲本 liao2345 相比，liao2345/o2-1 的赖氨酸浓度相似，而 liao2345/o2-2 的赖氨酸浓度增加了一倍。SDS-PAGE 分析表明，liao2345/o2-1 的醇溶蛋白比例与 liao2345 相同，而 liao2345/o2-2 的醇溶蛋白浓度明显降低。序列和转录丰度分析表明，两个 o2 NIL 的 CDS 均来自 CA339，但它们具有不同的启动子。由于 rbg 转座子插入到 liao2345/o2-2 的 TATA 盒和起始密码子之间，liao2345/o2-2 的 O2 转录物大部分被抑制。我们得出结论，o2 NIL 构建过程中不同的交叉互换模式导致了两个 o2 NIL 的不同籽粒表型。我们推测，liao2345/o2-1 返回到野生型是由于在导入和回交过程中与野生型 liao2345 启动子的重组。供体（CA339）的 o2 突变基因由于 O2 表达水平低而成为无效突变体。然而，其 CDS 可能编码具有正常功能的蛋白质，可维持玉米胚乳中醇溶蛋白的正常积累。
