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中文摘要：
溶菌酶（lysozyme）作为抗菌肽重要一员,在昆虫先天免疫系统中起着重要作用。本文通过家蝇EST序列筛选并结合RACE技术克隆了家蝇Musca domestica的lysozyme2基因（Md-lysozyme 2,MdL2）全长516bp的cDNA序列。MdL2包含429bp的开放阅读框,编码142个氨基酸残基,推导的氨基酸序列N端包括20个氨基酸残基的信号肽,成熟肽由122个氨基酸残基组成,理论分子量为13.89kD,理论等电点为6.45,有8个半胱氨酸组成4对分子内二硫键。实时荧光定量PCR结果显示大肠杆菌和金黄色葡萄球菌分别刺激后,家蝇幼虫体内MdL2基因的表达模式发生了非常相似的变化。在刺激3～12h表达下调,12～24h出现一个短暂的恢复,在刺激24h后表达又开始下调。不同组织定量PCR结果表明MdL2表达量在肠和血细胞中较高,而在表皮和脂肪体中较低。MdL2成熟肽编码序列被克隆入pET-DsbA表达载体,并转化大肠杆菌后得到高效表达。
英文摘要：
As an important member of the antimicrobial peptide family,lysozyme plays a key role in insect innate immune system.A full-length cDNA（516 bp） of lysozyme 2 gene was cloned from housefly（Musca domestica） by RACE based on the EST information.The cDNA contains a 429 bp open reading frame（ORF） encoding 142 amino acid residues and containing a putative signal peptide of 20 amino acids.The mature protein has 120 amino acids（21～42 aa） including 8 structural cysteine residues which form 4 pairs of disulfide bonds,with a molecular mass of about 13.89 kD and pI value of 6.45.The result of real-time quantitative PCR indicated that MdL2 gene expression patterns changed in very similar patterns in Musca domestica larva post injection by Escherichia coli and Staphylococcus aureus.Expression level declined from 3 to 12 h post-injection,and recovered to normal from 12 to 24 h post-injection,then started to drop again.MdL2 expressions in different tissues show that MdL2 was highly expressed in intestine and hemocyte,but expressed at relatively low levels in epidermis and fat body.MdL2 was cloned into a prokaryotic expression vector pET-DsbA and expressed efficiently in E.coli BL21（DE3）.
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