大豆腺苷酸激酶基因GmADK的克隆与表达分析

腺苷酸激酶(adenylate kinase，ADK)催化ATP+AMP⇔2ADP的可逆反应，是维持细胞能量动态平衡的关键酶，在植物中参与调节生长发育和逆境应答等过程。目前有关大豆ADK的研究还未见报道。本文通过RT-PCR方法，从耐盐大豆品种南农1138-2的叶中克隆到一个腺苷酸激酶基因，命名为GmADK。GmADK的编码区序列(coding DNA sequence, CDS)长804 bp，编码267个氨基酸。预测其蛋白结构含有典型的腺苷酸激酶功能域ATP-AMP(Ap5A)结合位点和AMP结合位点。蛋白序列比对和进化树分析表明，大豆与菜豆(Phaseolus vulgaris)和蒺藜苜蓿(Medicago truncatula)中的ADK序列相似性最高，亲缘关系最近。组织表达显示GmADK基因的表达量在大豆叶和根中高于茎中。荧光定量PCR分析表明GmADK的表达受盐胁迫的调节，且在耐盐(南农1138-2)和盐敏感(科丰1号)品种间存在差异。在叶和根中, 200 mmol L^–1 NaCl处理6、12和24 h后，GmADK的表达量在盐敏感品种中比未处理对照有所降低，但在耐盐品种中却比未处理对照升高，因此推测GmADK可能参与大豆对盐胁迫的响应。

Cloning and Expression Analysis of an Adenylate Kinase Gene GmADK in Soybean

Adenylate kinase (ADK) is an important enzyme in cellular energy homeostasis, which catalyzes the interconversion of adenine nucleotides, ATP+AMP⇔2ADP, and is involved in many processes including plant growth, development and response to abiotic stresses. To date, there has no report on cloning and expression analysis of soybean ADK gene yet. In this study, a soybean ADK gene was cloned from the leaves of a salt tolerant cultivar NN1138-2 using RT-PCR, and was designated as GmADK. The coding sequence (CDS) of this cloned ADK gene is 804 bp in length, encoding a polypeptide of 267 amino acids. Its protein was predicted to be located in plastids, containing a typical ATP-AMP (Ap5A) binding site and an AMP binding site. Multiple sequence alignments and phytogenetic analysis of ADK proteins showed GmADK is most similar with ADK from Phaseolus vulgarisand Medicago truncatula. Tissue expression pattern of GmADK showed its mRNA was more abundant in soybean roots and leaves than in stems. Quantitative RT-PCR showed a differential expression pattern of GmADK in response to salt between salt tolerant (NN1138-2) and sensitive (Kefeng-1) soybean genotypes. Compared with untreated plants, the expression of GmADK in both leaves and roots was repressed after 6, 12, and 24 hours of 200 mmol L^–1 NaCl treatment in the salt sensitive cultivar, but induced in the salt tolerant cultivar, indicating that GmADK might be involved in soybean response to salt stress.