鹿志伟 侯晓婉 杨子平 张燕梅 李俊峰 周文钊

摘  要  单子叶甘露糖结合植物凝集素基因能够有效抑制具有刺吸式口器的同翅目害虫的生长与繁殖，它被证实在掌叶半夏、小麦、烟草、棉花等单子叶和双子叶植物中均具有较好的抗虫效果，但其在剑麻中的功能尚缺乏深入研究。本研究以剑麻为材料，利用RT-PCR技术成功克隆剑麻lectin基因，命名为AsLEC，并对其进行生物信息学分析。结果表明：该基因CDS序列全长为561 bp，編码186个氨基酸，蛋白质分子量预测为19.97 ku，理论等电点为4.96，为疏水性蛋白;同源氨基酸序列比对结果表明剑麻lectin基因与火烧兰、雪花莲、君子兰与菠萝等植物的单子叶甘露糖结合凝集素基因同源性较高，氨基酸匹配度达到48%以上，系统进化树分析表明AsLEC基因与火烧兰亲缘关系更近;对AsLEC基因进行功能结构域分析表明其具有B-lectin基因家族典型特征，属于B-lectin基因家族成员;功能预测发现其具有一段33个氨基酸残基的信号肽，同时在N端第5～27位置处存在一个跨膜α螺旋，表明AsLEC蛋白为分泌型蛋白质，这与B-lectin蛋白的功能特点相吻合;AsLEC蛋白二级结构包含11个β折叠，3个α螺旋;亚细胞定位预测该基因很有可能定位在细胞膜上。剑麻AsLEC基因的克隆对于研究其在剑麻中的抗虫功能具有重要意义，同时丰富了单子叶植物中植物凝集素的相关研究成果。

关键词  剑麻;单子叶甘露糖结合植物凝集素;lectin

中图分类号  S563.8      文献标识码  A

Abstract  The monocotyl mannose-binding plant lectin gene can effectively inhibit the growth and reproduction of Homoptera pests with sucking mouthparts， and it was verified that those genes represent better resistance against insects in monocotyledonous and dicotyledonous plants such as Pinellia ternata， Triticum aestivum L.， Nicotiana tabacum L.， Gossypium spp. However， its function in sisal is still lack of in-depth study. In this study， sisal was used as materials， and the sisal lectin gene was successfully cloned by RT-PCR and named as AsLEC. Bioinformatics analysis were conducted. The results showed that the full-length CDS of AsLEC gene was 561 bp， encoding 186 amino acids. The protein molecular weight was predicted to be 19.97 ku， and the theoretical isoelectric point was 5.01， which was a hydrophobic protein. The results of homologous amino acid sequence alignment indicated that sisal lectin gene had high homology with the superfamily of monocotyl mannose-binding lectin gene in Epipactis helleborine， Leucojum vernum， Cliviaminiata， Ananas comosus （Linn.） Merr. and so on. The amino acid matching degree was more than 48%. The phylogenetic tree analysis indicated that AsLEC gene was more closely related to the lectin genes in Epipactis helleborine. Functional domain analysis of AsLEC gene suggested that it had the typical characteristics of the B-lectin gene family and belongs to the B-lectin gene family. A signal peptide of 33 amino acid residues was predicted in AsLEC protein， and there was a transmembrane alpha helix at the position from the 5th amino acid to the 27th amino acid in the N-terminus， which implyed that AsLEC protein was a secreted protein and was consistent with the functional characteristics of B-lectin protein. AsLEC protein secondary structure contained 11β-sheets， 3α-helices. Subcellular localization prediction showed that AsLEC gene was likely positioned on the cell membrane. Thecloning of sisal AsLEC gene was of great significance for studying its anti-insect function in sisal， and enriches the related research of plant lectin in monocotyledon.

Keywords  Agave sisalana; monocotyl mannose-binding plant lectin; lectin

DOI  10.3969/j.issn.1000-2561.2019.06.013

新菠萝灰粉蚧（Dysmicoccus neobrevipes，Beardsley）属于同翅目（Homoptera）、粉蚧科（Pseudococcidae）、洁粉蚧属（Dysmicoccus），其主要分布在热带、亚热带地区，如夏威夷、斐济、牙买加、马来群岛、墨西哥、密克罗尼西亚、菲律宾以及中国台湾等国家和地区。……