cDNA cloning,functional expression and antifungal activities of a dimeric plant defensin SPE10 from <Emphasis Type="Italic">Pachyrrhizus erosus</Emphasis> seeds

SPE10 is an antifungal protein isolated from the seeds of Pachyrrhizus erosus. cDNA encoding a 47 amino acid peptide was cloned by RT-PCR and the gene sequence proved SPE10 to be a new member of plant defensin family. The synthetic cDNA with codons preferred in yeast was cloned into the pPIC9 plasmid directly in-frame with the secretion signal -mating factor, and highly expressed in methylotrophic Pichia pastoris. Activity assays showed the recombinant SPE10 inhibited specifically the growth of several pathogenic fungi as native SPE10. Circular dichroism and fluorescence spectroscopy analysis indicated that the native and recombinant protein should have same folding, though there are eight cystein residues in the sequence. Several evidence suggested SPE10 should be the first dimeric plant defensin reported so far.Nucleotide sequence data reported in this paper are available in the DDBJ/EMBL/GenBank database under accession number AY679170     

豆薯种子中两种蛋白质的分离纯化及其性质研究

豆薯（Ｐａｃｈｙｒｒｈｉｚｕｓｅｒｏｓｕｓ）种子经磷酸盐缓冲液抽提,Ｓ－ＳｅｐｈａｒｏｓｅＦａｓｔＦｌｏｗ柱,ＤＥ－５２纤维素柱和ＳｅｐｈａｄｅｘＧ－７５柱层析,提取出两种高纯度的蛋白成分,命名为ＰａｃｈｙｒｉｎⅠ和Ⅱ．ＳＤＳ－ＰＡＧＥ测得其分子量分别为３３ｋＤ和１４．５ｋＤ,但ＨＰＬＣ分子筛的结果显示ＰａｃｈｙｒｉｎⅡ的分子量为２８ｋＤ,无论在还原条件下,还是在非还原条件下,ＰａｃｈｙｒｉｎⅡ电泳的结果都完全相同,表明该蛋白的亚基不是以二硫键相连。两种蛋白的等电点分别为４．５和６．５．用酸解法测定了它们的氨基酸组成。在无细胞体系中,它们对蛋白合成有较弱的抑制活性,显示它们可能是核糖体失活蛋白（ＲＩＰｓ）家族中的新成员。     

豆薯种籽中一种抗真菌蛋白PaAFP的分离纯化和部分特性研究

各类植物由于缺少自身免疫系统的支持,因而必须依赖于其它机制来抵御外来微生物的入侵．其中的一种重要机制就是通过合成体内各类能抑制微生物生长的蛋白质来完成的［１］．已报道从植物中分离出多种不同的抗真菌蛋白．广为研究的是几丁质酶和β－１,３－葡聚糖酶,认为它们在植物对真菌病的抗性中起重要作用［２,３］;核糖体失活蛋白（ＲＩＰｓ）和一类富含半胱氨酸的碱性多肽Ｔｈｉｏｎｉｎｓ也显示有非专一的抗真菌活性［４,５］．但仍有一些蛋白质,体外表现强烈的抗真菌活性,却不属于以上范围［６,７］．本文报道了豆薯种籽中一…     

不同贮藏条件下豆薯种子的脂质过氧化研究

豆薯(Pachyrhizuus erosus(L.)Urban)种子分别在4℃、15℃相对湿度(RH)45%及室温开放条件下贮藏,或新鲜种子进行超干处理将其含水量降到6.43%后密封贮存,以探讨种子劣变的生理机制.结果表明,豆薯种子在室温开放贮藏12个月的发芽率降至50%,24个月后发芽力完全丧失.4℃下贮藏豆薯种子最有效,贮藏后种子的电导率和紫外吸收值较低,过氧化氢酶、过氧化物酶和超氧化物歧化酶活性仍保持较高水平,种子萌发时释放出的挥发性醛也较低.适当的超干处理效果与15℃45%RH相似,但不及4℃的.     

豆薯种子中的杀虫成分及其毒力测定

为明确豆薯Pachyrrhizus erosus (L.) Urban种子中的杀虫成分及其杀虫毒力,以白纹伊蚊Aedes albopictus 4龄幼虫为目标昆虫,在活性跟踪的基础上,通过多种色谱技术和核磁共振技术分离鉴定其化学成分结构,并通过浸渍法、点滴、夹毒叶碟法等测定各化合物对白纹伊蚊幼虫、棉蚜Aphis gossypii无翅成蚜、甘薯天蛾Herse convolvuli幼虫和小菜蛾Plutella xylostella幼虫的杀虫活性和作用方式。结果表明,从豆薯种子中共分离、鉴定了14个化合物,即12a-hydoxyrotenone,pachyrrhizine,12a-hydoxypachyrrhizone,12α-dehydropachyrrizone,α-naphthoflavone,7-methoxyflavanone,12α-hydroxydolineone,6-methoxyflavone,4′-hydoxyflavanone,quercetin dihydate,5-methoxyflavone,7-hydroxyflavone,3′-hydoxyflavanone和3-hydroxyflavone。上述化合物中的前7个化合物对白纹伊蚊4龄幼虫具有毒杀活性, 处理24 h的LC₅₀分别是25.0,51.1,196.2,48.4,98.9,107.2 和15.6 mg/L;前6个化合物对棉蚜成虫24 h 的毒力分别为1.5,10.9,80.7,8,32.1和112.8 mg/L;前5个化合物以及12α-hydroxydolineone对甘薯天蛾3龄幼虫有毒杀活性;12α-hydoxyrotenone对3龄小菜蛾幼虫24 h的胃毒毒力LD₅₀为17.3 μg/头,7-methoxyflavanone对该虫仅表现出较弱的生长发育抑制活性。首次从豆薯种子中分离得9个化合物,即6-methoxyflavone,4′-hydoxyflavanone,quercetin dihydate,α-naphthoflavone,7-methoxyflavanone,5-methoxyflavone,7-hydroxyflavone,3′-hydoxyflavanone和3-hydroxyflavone;豆薯种子含有7个活性成分,其作用方式因目标昆虫不同而不同,其主要杀虫成分不是鱼藤酮而是12α-羟基鱼藤酮。     

Assimilate partitioning in Pachyrhizus erosus tubers under short days

The distribution of ¹⁴C products in yam bean ( Pachyrhizus erosus ) plants was measured as a function of time after a ¹⁴CO₂ pulse-chase experiment under short-day conditions. The ¹⁴C declined by more than 70% in leaves during the first two hours, indicating that they actively exported carbon. Tubers were strong storage sinks for carbon and accumulated more than 80% of the total incorporated ¹⁴C after a 72-h chase. The data show that, although sucrose represented about 21% of the tuber non-structural carbohydate, i.e. 15% of the tuber dry weight, ¹⁴C did not accumulate in sucrose but in glucose, fructose and starch. The data indicate the existence of a separate sucrose pool which is affected only very slowly by recent assimilates. As a result, recent photosynthates, temporarily stored, may contribute to the reservoir of carbon available for nitrogen fixation. The data also suggest the existence of two distinct pools of amino acids in the tuber, one utilized mostly for protein synthesis and the other probably stored in the vacuole.     

Effects of bark beetle pheromones on the attraction of Monochamus alternatus to pine volatiles

Abstract We evaluated the attraction of Monochamus alternatus Hope (Coleoptera: Cerambycidae), Dryocoetes luteus Blandford and Orthotomicus erosus Wollaston (Coleoptera: Curculionidae: Scolytinae) to multiple-funnel traps baited with the pine volatiles, ethanol and (+)-α-pinene and the bark beetle pheromones, ipsenol and ipsdienol. M. alternatus were attracted to traps baited with ethanol and (+)-α-pinene but not those baited with ipsdienol and ipsenol. Ipsdienol and ipsenol decreased catches of M. alternatus in traps baited with ethanol and (+)-α-pinene. Traps baited with either binary combinations of ethanol and (+)-α-pinene or ipsdienol and ipsenol were attractive to D. luteus and O. erosus. The addition of ipsenol and ipsdienol to traps baited with ethanol and (+)-α-pinene synergized attraction of O. erosus but not D. luteus.