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为研究水稻蜡质基因（ｗａｘｙ）５＇上游调控区中存在的顺式作用元件，我们将水稻ｗａｘｙ基因翻译起始声、（ＡＴＧ）５＇上游３．４ｋｂ（－２１１８～＋１２９１ｂＰ）片段经外切核酸酶ＥｘｏⅢ部分酶解，得到一系列５＇端缺失的片段。将这些缺失片段分别与ｇｕｓ基因编码区连接，构建成融合质粒，经ＰＥＧ介导引入水稻原生质体，２６℃培养４８ｈ后，定量测定ＧＵＳ酶活力，并以同时导入的由３５Ｓ启动子指导的荧光素酶（ＬＵＣ）基因表达的酶活力作为内对照。结果表明，ＧＵＳ酶活性随５’上游调控区长度的减少而逐渐减弱。由─８６１ｂｐ缺失至─６４０ｂＰ时，ｇｕｓ基因表达水平有较明显的降低，推测在该区域中可能存在一个顺式作用元件区。
In order to study the cis-acting elements present in the upstream regulatory region of 5 ’waxy rice, we cloned the 3.4kb (-2118 ~ +1291 bp) upstream of the 5’ upstream of the (ATG) Partial digestion with exon III gives a series of 5 ’deletion fragments. These missing fragments were respectively ligated with the coding region of gus gene to construct a fusion plasmid. After being introduced into rice protoplast by PEG, the enzyme activity of GUS was quantitatively determined after culturing at 26 ° C. for 48 h, and the fluorescence of 35S promoter guided simultaneously Enzyme (LUC) gene expression of enzyme activity as an internal control. The results showed that the enzyme activity of GUS gradually decreased with the decrease of 5 ’upstream regulatory region. From -861bp to -640bP, the expression level of gus gene was significantly reduced, suggesting that there may be a cis-acting element region in this region.