Winkler R G, Helentjaris T
Department of Plant Sciences, University of Arizona, Tucson 85721, USA.
Plant Cell. 1995 Aug;7(8):1307-17. doi: 10.1105/tpc.7.8.1307.
Gibberellins (GAs) are phytohormones required for normal growth and development in higher plants. The Dwarf3 (D3) gene of maize encodes an early step in the GA biosynthesis pathway. We transposon-tagged the D3 gene using Robertson's Mutator (Mu) and showed that the mutant allele d3.2::Mu8 is linked to a Mu8 element. The DNA flanking the Mu8 element was cloned and shown to be linked to the d3 locus by mapping in a high-resolution population developed by selecting for recombination between d3 and linked genetic markers. To establish unambiguously the identity of the cloned gene as D3, a second mutant allele of D3 (d3.4) was also cloned and characterized using the d3.2::Mu8 sequences as a probe. d3.4 was found to have a novel insertion element, named Sleepy, inserted into an exon. A third mutant allele, d3.1, which has the same size 3' restriction fragments as d3.4 but different 5' restriction fragments, was found to contain a Sleepy insertion at the same position as d3.4. On the basis of the pedigree, Sleepy insertion, and restriction map, d3.1 appears to represent a recombinational derivative of d3.4. The D3 gene encodes a predicted protein with significant sequence similarity to cytochrome P450 enzymes. Analysis of D3 mRNA showed that the D3 transcript is expressed in roots, developing leaves, the vegetative meristem, and suspension culture cells. We detected reduced D3 mRNA levels in the mutant allele d3.5.
赤霉素（GAs）是高等植物正常生长发育所需的植物激素。玉米的矮化3（D3）基因在赤霉素生物合成途径中编码早期步骤。我们使用罗伯逊氏转座子（Mu）对D3基因进行转座子标签定位，并表明突变等位基因d3.2::Mu8与一个Mu8元件相连。克隆了Mu8元件侧翼的DNA，并通过在由d3与连锁遗传标记之间的重组选择所构建的高分辨率群体中进行定位，表明其与d3位点相连。为了明确克隆基因就是D3，还克隆了D3的第二个突变等位基因（d3.4），并以d3.2::Mu8序列作为探针进行了表征。发现d3.4有一个名为Sleepy的新插入元件插入到一个外显子中。发现第三个突变等位基因d3.1与d3.4具有相同大小的3' 限制性片段，但5' 限制性片段不同，其在与d3.4相同的位置含有一个Sleepy插入。根据系谱、Sleepy插入和限制性图谱，d3.1似乎代表d3.4的重组衍生物。D3基因编码一种预测的蛋白质，与细胞色素P450酶具有显著的序列相似性。对D3 mRNA的分析表明，D3转录本在根、发育中的叶片、营养分生组织和悬浮培养细胞中表达。我们在突变等位基因d3.5中检测到D3 mRNA水平降低。
