原位杂交技术及其在果树研究中的应用

原位杂交技术是近年来快速发展起来的一门新技术,本文介绍了原位杂交技术的基本原理、方法及其发展前景,以及该技术与其它生物学技术相结合而形成的一些新技术。综述了这些技术在果树研究中的应用情况。     

百合杂种系间杂交障碍的克服及3种幼胚离体培养方法的比较研究

分别以麝香百合(Longiflorum,L)'White Fox'、亚洲百合(Asiatic,A)'Conneticut King'、东方百合(Oriental,O)'Sorbonne'及喇叭百合(Trumpet,T)'Royal Goden'为亲本,用切割柱头法进行杂交,采用离体培养技术对杂种幼胚进行抢救,并运用基因组原位杂交技术对所得杂种进行鉴定.结果显示,在所选的6个杂交组合中,切割柱头授粉能不同程度地克服除LL×TT之外的5个杂交组合的受精前障碍;胚、胚乳及胚珠3种外植体的离体培养萌发率分别为100%、21.5%和1%,所得植株占所得杂种的比例分别为9.6%、89.9%和0.5%;基因组原位杂交分析显示,所获植株均为真杂种.结果证明,胚、胚乳及胚珠培养是从幼胚获得百合杂种植株的高效途径,且能有效防止伪杂种混杂,胚乳培养是获得百合杂种一代的主要方式.     

甘菊CMO同源基因的分离与表达分析

利用半嵌套巢式PCR结合RACE技术从菊科植物甘菊[Chrysanthemum lavandulifolium(Fisch.ex Trautv.)Makino]中分离得到了一个长度为1 450 bp的片段.序列分析结果表明,其开放阅读框全长1 140bp,编码379个氨基酸残基;在GenBank 比对并进行系统进化分析可知,该片段为CMO同源基因,命名为ClCMO.利用不同胁迫处理进行分析发现,在非胁迫条件下ClCMO基因在甘萄茎、叶、花叶中均有表达信号,在根中没有表达;其可以响应干旱、高盐胁迫和脱落酸(ABA)的诱导,不响应冷热胁迫,并且其表达在水杨酸(SA)诱导下受抑制.这些结果表明,ClCMO基因是提高植物耐干旱、高盐能力的有效基因资源.     

寄生取食Cry1Ab杀虫蛋白的粘虫幼虫对伞裙追寄蝇寄生率及生长发育的影响

【Aim】 To demonstrate the effect of transgenic Bt-crop on the bionomics of the tachinid fly Exorista civilis Rondani, the parasitism rate and life history parameters of E. civilis developed from larvae of the oriental armyworm, Mythimna separata (Walker), fed on artificial diets with Cry1Ab toxin were investigated in laboratory. 【Methods】 M. separata larvae were fed on diets containing 0, 3.125, 6.25, 12.5 and 25 μg/g of Cry1Ab toxin, respectively, and used as the host for E. civilis. The parameters related to the parasitism and the growth and development of E. civilis on the host were observed and recorded. 【Results】 The results showed that the parasitism rate and fecundity of the fly on M. separata larvae feeding on diets containing various doses of Cry1Ab protein were insignificantly different (P≥0.05), suggesting that host selective behavior of the E. civilis is not affected by the dose of Bt-toxin in the caterpillar. The developmental period (egg-larva, pupa, adult oviposition and generation), emergence rate, sex ratio, and fecundity of E. civilis developed from M. separata larvae treated by various concentrations of Cry1Ab toxin were insignificantly different (P≥0.05). However, the survival rate of the maggots in the host feeding on diet containing 3.125 μg/g of Cry1Ab toxin was significantly greater than that of the control and the rest treatments (P<0.05), while insignificantly different among the remained 4 treatments (P≥0.05). The pupal weight significantly declined as the concentration of Cry1Ab toxin fed by their hosts increased (P<0.05). The pupal weight in the control group was 49.8 mg, while that in treatment groups of 6.25 and 25 μg/g Cry1Ab toxin was only 41.0 and 36.7 mg, respectively. 【Conclusion】 These results indicate that parasitizing host M. separata larvae fed with Cry1Ab toxin has no dramatically negative effect on parasitoid E. civilis. This study provides a basis for the evaluation of the risk of transgenic Bt-crops on the tachinid parasitoids in fields.     

新疆核桃早实特性及RAPD分析

用RAPD技术进行新疆核桃早实特性的分子标记研究,用180个10-mer随机引物分别扩增早实和晚实近等基因池DNA,筛选出5个多态性引物,结果只有引物OPG15（5’-ACT GGG ACT C-3’）在早实近等基因池及其个体中能重复扩增出一条约710bp的特异片段OPG15 710,而晚实近等基因池及其个体中无此特异片段,将OPG15 710克隆于pUCm-T载体,筛选阳性克隆用限制性内切酶Pst1消化,表明克隆片段大小正确。实步分析认为,OPG15 710可能是与核桃早实特性相关的分子标记,该标记仍在研究之中,该研究将为进一步克隆早实基因,实现早实优质,探索早实机制奠定基础。     

家蝇卵巢中的信息化学物质引诱雌蝇产卵的研究

产卵期家蝇MuscadomesticaL .卵巢中的信息化学物质诱引家蝇产卵。行为试验表明 ,雌蝇产卵时优先选择被成熟卵巢污染过的发酵麦麸 ,诱集的产卵百分率依次为 :发酵麦麸 + 1对卵巢 >湿麦麸+ 1对卵巢 >发酵麦麸 >湿麦麸 ;卵巢提取物同样对产卵雌蝇有诱集活力。产卵雌蝇卵巢正己烷、二氯甲烷以及乙酸乙酯的提取物均能诱集雌蝇产卵 ,诱集的产卵百分率依次为 :雌蝇卵巢正己烷提取物 >乙酸乙酯提取物 >二氯甲烷提取物。推测家蝇卵巢中的信息化学物质可能含有家蝇产卵聚集信息素的组分。     

蛋白质谷氨酰胺酶的发酵及初步分离和应用研究

研究了不同发酵条件对于产吲哚金黄杆菌（Chryseobacterium indologenes）生产蛋白质谷氨酰胺酶能力的影响,酶的分离和初步的应用。通过考察种子的生长曲线,发酵的温度、转速、摇瓶装液量,碳源和氮源,得出最大产酶能力的发酵条件为：30℃,200r/min,装液量25ml/250ml,蔗糖为碳源,多聚蛋白胨为氮源,发酵10~12h。初步分离研究表明在4倍超滤浓缩和4倍乙醇沉淀条件下,酶活力回收率均为最高,分别为84.99%和76.07%。该酶与酪蛋白37℃温育2h时,酪蛋白的脱酰胺度为41.03%,到24h后,脱酰胺度不再增加,并且酪蛋白的溶解性也有所增加。