2 REGULATORY GENES OF THE MAIZE ANTHOCYANIN PATHWAY ARE HOMOLOGOUS - ISOLATION OF B UTILIZING R GENOMIC SEQUENCES
2 REGULATORY GENES OF THE MAIZE ANTHOCYANIN PATHWAY ARE HOMOLOGOUS - ISOLATION OF B UTILIZING R GENOMIC SEQUENCES
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DOI：
10.1105/tpc.1.12.1175
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发表时间：
1989-12-01
期刊：
影响因子：
11.6
通讯作者：
TURKS, D
中科院分区：
文献类型：
--
作者：
CHANDLER, VL;RADICELLA, JP;TURKS, D
玉米的遗传学研究已经确定了几个调控基因，这些基因在发育过程中控制紫色花青素色素的组织特异性合成。两个这样的基因，R和B，在花青素合成的组织特异性和发育时间方面表现出广泛的等位基因多样性。先前的遗传学研究表明，某些B等位基因可以替代R的功能，在这些情况下，糊粉层中色素合成只需要两个基因座中的一个有功能的等位基因。此外，生化研究表明，这两个基因作用于相同的生物合成途径，这表明这两个基因在功能上是重复的。在本报告中，我们描述了DNA杂交实验，这些实验表明B和R在功能上的重复性质反映在这两个基因之间的DNA序列相似性上。我们利用这种同源性，使用R基因组序列来克隆B。我们采用了两种不同的策略，分离出了两个基因组克隆，一个是与W23自交系中的b等位基因相连的2.5千碱基对的BgIII片段，另一个是与CM37品系中的B等位基因相连的1.0千碱基对的HindIII片段。对B中的几个独立的转座因子插入突变和回复突变体的检测表明，我们的克隆识别有功能的B基因。代表整个B - Peru等位基因的基因组克隆被分离出来，并绘制了详细的限制性内切酶图谱。利用这些克隆，我们在含有B - I或B - Peru等位基因的植株的苞叶中鉴定出一种2.2千碱基对的mRNA，但在含有b等位基因的植株中未检测到B mRNA。在色素深的B - I苞叶组织中，该转录本的含量比色素浅的B - Peru苞叶组织中至少高100倍。有功能的B等位基因在苞叶组织中的表达与该途径中两个结构基因A1和Bz1的mRNA水平的协同增加相关，这与B作为调控基因的假定作用是一致的。
Genetic studies in maize have identified several regulatory genes that control the tissue-specific synthesis of the purple anthocyanin pigments during development. Two such genes, R and B, exhibit extensive allelic diversity with respect to the tissue specificity and developmental timing of anthocyanin synthesis. Previous genetic studies demonstrated that certain B alleles can substitute for R function, and in these cases only one functional allele at either locus is required for pigment synthesis in the aleurone. In addition, biochemical studies have shown that both genes act on the same biosynthetic pathway, suggesting that the genes are functionally duplicate. In this report we describe DNA hybridization experiments that demonstrate that the functionally duplicate nature of B and R is reflected in DNA sequence similarity between the two genes. We took advantage of this homology and used the R genomic sequences to clone B. Two different strategies were pursued and two genomic clones isolated, a 2.5-kilobase BgIII fragment linked to the b allele in W23 inbred stocks and a 1.0-kilobase HindIII fragment linked to the B allele in CM37 stocks. Examination of several independent transposable element insertion mutations in B and revertant derivatives demonstrated that our clones recognize the function B gene. Genomic clones representing the entire B-Peru allele were isolated, and a detailed restriction map was prepared. Using these clones we have identified a 2.2-kilobase mRNA in husks from plants containing either B-I or B-Peru alleles, but no B mRNA was detected in plants containing a b allele. The transcript is at least 100 times more abundant in strongly pigmented B-I husks than in weakly pigmented B-Peru husk tissue. Expression of functional B alleles in husk tissue correlates with the coordinate increase in mRNA levels of two structural genes of the pathway, A1 and Bz1, consistent with the postulated role of B as a regulatory gene.