LIU Jiang,CHEN Pei,XING Guang-nan,et al.Cloning of the Soybean Chalcone Reductase Gene GmCHR and Construction of Its Plant Expression Vector[J].Soybean Science,2013,32(02):139-142.[doi:10.3969/j.issn.1000-9841.2013.02.001]
大豆查尔酮还原酶基因GmCHR的克隆与植物表达载体构建
- Title:
- Cloning of the Soybean Chalcone Reductase Gene GmCHR and Construction of Its Plant Expression Vector
- Keywords:
- Soybean; Isoflavone; Chalcone reductase ; Gene expression
- 文献标志码:
- A
- 摘要:
- 黄酮类化合物在植物中参与过滤紫外线、固氮和花色形成等过程， 异黄酮 对人体有抗氧化、预防乳腺癌等的保健作用。 查尔酮还原酶（ chalcone reductase ， CHR ）是植物中参与黄酮 类化合物 代谢的重要酶。 克隆大豆查尔酮还原酶基因并构建植物表达载体，有助于进一步研究其功能和异黄酮的代谢过程。 采用 RT-PCR 方法，从栽培大豆 （ Glycine max ） 南农 1138-2 中，克隆得到了第 14 号染色体上的一个编码大豆查尔酮还原酶（ chalcone reductase ， CHR ）的基因，命名为 GmCHR 。该基因含有 948 bp 长的编码区序列 (Coding DNA Sequence,CDS), 编码 315 个氨基酸。预测其蛋白质分子量为 35.5 KDa ，等电点为 6.32 。与其他豆科植物中的查尔酮还原酶相比， GmCHR 蛋白序列与 葛藤（ Puerariae montana ） CHR 的 相似性最高，达 94% 。组织表达分析表明，在自然生长条件下 GmCHR 在叶中的表达量最大；其次是种子；在花和茎中相同；在根中的表达量最小。利用 Gateway 方法获得植物过表达载体 pMDC83-GmCHR ，经检测表明过表达载体已成功转化农杆菌 EHA105 ，为今后进一步了解 GmCHR 在大豆异黄酮代谢过程中的功能提供材料基础。
- Abstract:
- Flavonoids are involved in UV filtration, symbiotic nitrogen fixation and floral pigmentation in plants. Isoflavones have potential effects on human health, such as antioxidant activity, preventing breast cancer and other cancers. Chalcone reductase (CHR) is an important enzyme involved in flavonoid biosynthesis. Cloning of soybean chalcone reductase and construction of its plant expression vector would help study its function and the phenylpropanoid pathway. A gene encoding CHR on chromosome 14 was cloned from the cultivated soybean （Glycine max (L.) Merr.）cultivar Nannong 1138-2 using RT-PCR, and was designated as GmCHR. This gene contains a coding DNA sequence (CDS) of 948 bp, and the corresponding protein consists of 315 amino acids. The protein is estimated to have a molecular weight of 35.5 kDa and isoelectric point of 6.32. Comparing with the amino acid sequence of CHR from other legume species, GmCHR has a highest similarity of 94% with the CHR from Puerariae Montana. The mRNA abundance of GmCHR was highest in leaves, followed by seeds, flowers, stems and roots under normal growth conditions. A plant over-expression vector of pMDC83-GmCHR was constructed by Gateway technology, and was transferred into Agrobacterium EHA105, which would provide the opportunity to study the function of GmCHR in soybean flavonoid biosynthesis.
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备注/Memo
?国家重点基础研究发展计划(973计划)项目(2009CB1184，2011CB1093)；国家自然科学基金项目(31071442)；江苏省优势学科建设工程专项和国家重点实验室自主课题；现代农业产业技术体系建设专项资金（CARS-04）；农业部大豆生物学与遗传育种创新团队。