Construction and Identification of Yeast Surface Display Libraries of Ovis aries DRA Gene Exon 2

The specific primers were designed according to Ovis aries DRA gene sequence deposited in GenBank and the multiple cloning site of the plasmid pYD1,which was a vector used for protein surface display on Saccharomyces cerevisiae.The gene encoding DRA was amplified by PCR using the genomic RNA of Ovis aries.The 762 bp fragment was cloned and released in GenBank and registration number was KR422362.The PCR product was inserted into the yeast surface display plasmid vector pYD1 by double enzyme digestion.It was indicated that DRA gene was successfully integrated into the genome.Dot mutation was made at both ends of exon 2 in DRA gene for making restriction enzyme cutting site and design the exon 2 specific primers according to mutated Ovis aries DRA gene sequence.Sequenced exon 2 amplification products based on DNA pooling of sheep large sample template was analyzed the polymorphic loci.The polymorphic exon 2 246 bp fragment was obtained by double enzyme digestion and connected to surface display restructuring mutation carriers pYD1-DRA by the same double enzyme digestion,and then we successfully constructed yeast surface display libraries.We transformed it into Saccharomyces cerevisiae EBY100 cell.Yeast monoclone was identified by PCR amplification and sequencing,and we confirmed that DRA gene had been integrated into Saccharomyces cerevisiae genome.After galactose induced,it was detected that DRA gene library had been successfully demonstrated on the yeast cell surface under the fluorescence microscope by immunofluorescence method.     

蒜和葱上二斑叶螨的发生为害及其抗药性监测

二斑叶螨寄主植物范围极广, 但其对百合科蔬菜造成危害的研究报道极少?本研究采集了来自北京海淀百合科蔬菜蒜和葱上的2个二斑叶螨种群, 生物测定结果显示, 2个种群对阿维菌素呈现出极高水平抗性, 对哒螨灵为中等以上抗性(LC50>5 000 mg/L), 对新型药剂联苯肼酯?虫螨腈和腈吡螨酯也表现出中等到高水平抗性?抗性基因突变频率检测发现, 与阿维菌素抗性相关基因G314D和G326E位点以及和哒螨灵抗性相关基因H92R位点的突变频率均高达100%, 为纯合抗性种群?因此, 本研究所测试的2个二斑叶螨种群对阿维菌素和哒螨灵单剂的敏感性极低, 需慎重选用这2种药剂, 而3种新型杀螨剂联苯肼酯?虫螨腈和腈吡螨酯在田间需轮换使用?     

Highly recurrent BRAF p.V595E mutation in canine papillary oral squamous cell carcinoma

Oral squamous cell carcinoma (OSCC) is the most common oral epithelial malignancy in dogs. It exhibits locally aggressive biological behaviour with the potential to metastasize, and a reported 1-year survival rate of 0% when left untreated. Expression studies suggest that aberrant MAPK signalling plays a key role in canine OSCC tumorigenesis, which is consistent with BRAF and HRAS MAPK-activating mutations reported in some tumours. Several morphological subtypes of canine OSCC have been described, with papillary, conventional, and basaloid as the most common patterns. We hypothesized that mutational differences may underlie these phenotypic variations. In this study, targeted Sanger sequencing and restriction fragment length polymorphism assays demonstrate that up to 85.7% of canine papillary OSCC (n = 14) harbour a BRAF p.V595E mutation. Assessment of neoplastic epithelial cell proliferation using Ki67 immunolabelling (n = 10) confirmed a relatively high proliferation activity, consistent with their known aggressive clinical behaviour. These findings underscore a consistent genetic feature of canine papillary OSCC and provide a basis for the development of novel diagnostic and targeted therapeutic approaches that can improve the quality of veterinary care.     

大豆化学诱变育种及其规律的研究

应用化学诱变剂处理大豆种子,对M_1代植株的生育及形态均有强烈的抑制作用.M_1代所产生的形态变异主要是由于化学物质扰乱了植株生理而产生生理损伤所致.在M_2代植株各性状中以分枝数、株高、单株荚数的遗传变异系数较大,说明这些性状选择的潜力是大的,通过选择容易达到预期目标.诱变M_3代绝大多数性状仍在分离.加强选择,能选出早熟、大粒、农艺性状好、丰产的新品系.     

5-氮胞苷对籼稻成熟胚愈伤组织生长和再分化的影响及诱变效应

用5-氮胞苷(5-azaC)处理籼型三系杂交水稻恢复系“明恢63”、“密阳46”和“特青2号”成熟胚诱导产生的愈伤组织,当5-azaC浓度为0.1mmol/L时,愈伤组织的增殖明显受抑;当5-azaC浓度达1.0mmol/L时,愈伤组织增殖率下降50%以上;5-azaC浓度提高到1.5mmol/L时,愈伤组织增殖率仅及对照的20%。分化阶段愈伤组织对5-azaC更敏感,当5-azaC的浓度为0.02mmol/L时,愈伤组织的再分化率明显下降,且随5-azaC浓度的增加,再分化率也显著下降,当5-azaC的浓度为0.2mmol/L时,各试验品种的愈伤组织的再分化力完全受抑,不能分化出幼苗。经5-azaC处理的愈伤组织再分化力也下降,但经预分化培养后分化力可恢复到较高水平。经5-azaC处理的愈伤组织再生植株产生了性状变异,出现了白化苗、矮秆、迟熟和早熟等变异体,当代的突变率为3.03%~5.31%。突变性状表现为单基因隐性突变,可稳定遗传。     

水稻不育系培矮64S的空间诱变效应及后代的SSR分析

对经过卫星搭载的培矮64S种子及后代进行了有关性状研究和SSR标记分析,结果表明:培矮64S空间诱变后代SP1代发芽率、存苗率、株高、抽穗期、株叶型的性状与对照相比没有明显变化;但SP2代在株高、粒型和育性等性状上发生了变异;从SP3代中获得柱头比培矮64S明显增大的变异株系,这对于改良培矮64S的异交结实率和提高制种产量具有重要意义。此外,对10个SP3变异株进行SSR分析结果表明,扩增片段数目变化约占55.21%,扩增片段分子量变化约占44.79%,空间诱变引起的变异可能是以DNA缺失-重复为主。SSR座位的变异频率平均为23.33%,变异座位在水稻基因组中是随机分布的。     

花生太空诱变育种研究初报

2003年利用返回式卫星搭载花生品种粤油7号种子,回收返地后种植。田间试验结果显示,SP1代未出现显著的变异,但在SP2和SP3中发现荚果大小、形状和种皮颜色等变异。太空诱变为开发新的花生种质和有用的基因提供了有效的方法。     

‘越心’草莓组培突变体着色差异的分子机理初探

初步探讨了在‘越心’草莓（Fragaria × ananassa‘Yuexin’）茎尖离体培养再生植株中发现的稳定变异突变体着色差异的分子机理。分析结果显示,突变体与‘越心’在植株形态、始花期、始果期、始熟期、单果质量、平均株产、果形、风味等方面均无明显差异;而突变体外果皮色泽较淡,L*值较高,a*值较低,H色度角值较大,颜色指数CIRG显著小,且果肉不着色（‘越心’果肉红色）。突变体果实总花青苷含量为‘越心’的13.2%,其中以天竺葵素–3–O–葡萄糖苷（Pg3G）含量差异最大,突变体Pg3G含量仅为‘越心’的12.9%。花青苷合成途径结构基因FaF3H、FaCHS、FaDFR、FaANS、FaCHI、Fa3GT的表达水平在突变体中显著降低,这可能是突变体成熟果实花青苷水平显著低于‘越心’的直接原因。FaMYB10和FaMYB1在突变体中的表达水平显著下降,且FaMYB10在几个主成分中的特征向量绝对值均显著较高,进一步证实了MYB10是草莓成熟过程中花青苷合成的类黄酮或苯丙烷类化合物途径主要调节因子之一。提取叶片基因组DNA进行基于KASP标记的基因分型结果显示,74对KASP引物在‘越心’草莓及其突变体中的基因分型不存在差异。本结果初步证实突变导致参与调控花青苷合成途径的关键基因表达发生显著下调从而使花青苷积累显著减少,果实着色变淡。