Function of a short spacer between rice EPSP synthase and rps20 genes.

我们将在水稻愈伤组织中表达的片段序列映射到水稻蜡质位点周围328 kb BAC的物理图谱上。在杂交片段中，我们试图表征位于蜡质下游约45 kb区域的强杂交片段。这些片段含有2个内控基因，分别编码5-烯醇丙酮酰莽草酸-3-磷酸合成酶（EPSPs）和核糖体蛋白小亚基20 （rps20）。有趣的是，这些管家基因是相邻的，只有一个303 bp的间隔，并以尾对尾的方向定向。通过对愈伤组织cDNA文库的筛选，获得了两个基因的近全长cDNA克隆。EPSPs基因克隆具有1683 bp的开放阅读框，编码561个氨基酸（aa）的多肽，这是该方法在水稻中首次发现的表达基因。EPSPs是莽草酸途径的关键酶，存在于高等植物的质体中。草甘膦是一种非选择性除草剂，这种酶更容易受到草甘膦的攻击。水稻rps20肽的基因已被分离出来，我们的rps2O cDNA克隆与该肽的序列完全匹配。利用从粳稻和籼稻品系的根、苗、颖花和花药中提取的总rna进行了Northern blotting分析。EPSPs基因的2.2 kb转录本在根、苗和小穗的RNA样本中很容易检测到，但在花药的RNA样本中检测不到。我们检测的所有组织中都检测到一个0.75 kb的rps20转录本，尽管这两个水稻品系的根中的表达都非常弱。我们对这两个基因的转录本进行了一定程度的检查。这些转录本超越了间隔；另一个基因的表达可能被来自相反基因的反义RNA修饰，产生双链RNA，这可能在转录后基因沉默机制中触发RNA降解。这里发现的短间隔在高等植物的核基因组织中是不常见的。该片段可能对植物核基因组中间隔物的扩增或收缩具有进化意义，并可能在相反基因转录引起的基因调控中发挥作用。少

We mapped the fragments with sequence (s) expressed in the rice callus onto a physical map for a 328-kb BAC contig around the rice waxy locus. Among the hybridized fragments, we attempted to characterize strongly hybridized fragments located in a region about 45 kb downstream of waxy. The fragments were found to contain two housekeeping genes encoding 5-enolpyruvylshikimate-3-phosphate synthase (EPSPs) and ribosomal protein small subunit 20 (rps20). Interestingly, these housekeeping genes are adjoined, with only a 303-bp spacer, and are directed in a tail-to-tail orientation. Nearly-full-length cDNA clones of the two genes were obtained by screening the callus cDNA library. A clone for the EPSPs gene possessed an open reading frame of 1683 bp encoding a deduced 561-amino-acid (aa) polypeptide that is the first expressed gene identified in rice using this method. EPSPs is a key enzynae on the shikimate pathway and is localized in the plastids of higher plants. This enzyme is known to be … More targeted by glyphosate, which has been employed as a nonselective herbicide. A gene for the rice rps20 peptide had previously been isolated, and our rps2O cDNA clone completely matched the registered sequence for this peptide. Northern blotting analyses were conducted using blots of total RNAs extracted from roots, seedlings, spikelets and anthers of Japonica and Indica rice strains. The 2.2-kb transcript of the EPSPs gene was easily detected in RNA samples from roots, seedlings and spikelets, but not the RNA from anthers. A 0.75-kb rps20 transcript was detected in all the tissues we examined, although the expression in roots from both rice strains was very weak. We have examined an extent of the transcript of either gene. These transcripts proceeded beyond the spacer ; the expression of the other gene might be modified by the antisense RNA from the opposite gene, resulting in double-stranded RNA, which might act as a trigger for RNA degradation in the mechanism called post-transcriptional gene silencing. The short spacer found here is uncommon in the nuclear gene organization in higher plants. This segment may have evolutionary implications about spacer expansion or contraction in the plant nuclear genome, and may play a role in gene regulation caused by the opposite gene transcript. Less