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摘要
以表皮生长因子（epidermal growth factor， EGF， Pfam PF00008.28）和激酶（pkinase， Pfam PF00069）保守域全蛋白序列为种子序列，鉴定了山荆子（Malus baccata） WAK-RLK（Wall-associated kinases- Reporter like kinase）家族成员，对其家族成员理化性质、进化关系、重复现象和表达模式等进行分析。结果表明，山荆子WAK基因家族包括27个成员，其氨基酸序列大小介于496～805，分子量介于55.45～89.61 kD，理论等电点介于5.15～8.75，主要分布于质膜；根据进化分析，将拟南芥、水稻、番茄、香蕉、苹果、山荆子6个物种的WAK-RLK分为8个亚组，山荆子WAK主要分布于亚组I、II、IV、V、VII和VIII。基因重复分析表明山荆子WAK成员扩增的主要原因是发生基因组范围的重复（wide-genome duplication， WGD）。启动子顺式作用元件（cis-acting regulatory element， cis-element）分析表明，该家族成员主要响应多种激素和逆境信号。PCR表达分析表明，WAK基因在山荆子不同组织中存在差异表达，且在苹果树腐烂病菌（Valsa mali）侵染后，MABA031460、MABA030455和MABA014609在前期显著上调表达。由此表明山荆子WAK家族成员发生扩增，响应逆境和腐烂病信号，MABA031460、MABA030455和MABA014609可作为抗腐烂病研究的候选基因。本研究结果为进一步抗病研究和功能分析提供了参考。Based on conserved protein sequence of EGF （ epidermal growth factor， EGF; Pfam PF00008.28 ） and Pkinase （ Pfam PF00069 ） domain， the WAKs of Malus baccata were identified at a genome-wide level. For all members， physicochemical parameters， evolutionary characteristics， duplication event and expressional patterns were investigated. In total， 27 WAKs were identified in the Malus baccata genome， with amino acid size， the molecular weight， and isoelectric point varying from 495～805 aa， 55.45～89.61 kD and 5.51～8.75， respectively. Among these， most of these were predicted on plasma membrane. Evolutionary analysis showed that WAKs from Arabidopsis thaliana， Oryza sativa， Solanum lycopersicum， Musa acuminata， Malus domestica and Malus baccata could be divided into 8 sub-groups， the members from Malus baccata were distributed in sub-group I， II， IV， V， VII and VIII. Gene duplication events indicated that WGD （Wide-genome duplication） was the main cause of duplication in Malus baccata WAKs and there were multiple cis-elements （cis-acting regulatory element， cis-element ） in promoter region in response to hormone and stress signal. Gene expression analysis showed that nine members of Malus baccata WAKs were showed diverse expression patterns in different tissues. MABA031460， MABA030455 and MABA014609 were significantly up-regulated when Malus baccata suspension cells were infected with Valsa mili （Vm） in the early stage. These differentially expressed genes could be used as candidate genes for disease resistance and further functional investigation.
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