一种利用微波炉加热快速提取细菌DNA用于PCR扩增的方法

利用微波加热原理,探索一种快速、简单、高效提取细菌DNA用于常规PCR快速检测的方法。在微波炉额定功率800 W条件下分别对大肠杆菌、金黄色葡萄球菌、单增李斯特氏菌、副溶血弧菌、沙门氏菌以及志贺氏菌6种常见食源性病原菌进行微波处理,离心取上清,获取细菌DNA,将样本DNA进行PCR扩增及琼脂糖凝胶电泳进行验证。并进一步采用微波加热方法针对6种病原菌最低检出浓度和混合提取6种病原菌DNA的PCR检测进行分析。6种病原菌在微波炉中加热40-130 s的上清DNA样本均能用于常规PCR扩增。针对检出浓度的分析表明,副溶血弧菌的最低检出浓度为103 cfu/ml,金黄色葡萄球菌为104 cfu/ml,肠出血性大肠杆菌为105 cfu/ml,肠炎沙门氏菌和福氏志贺氏菌的最低检出浓度为106 cfu/ml,针对单核细胞增生李斯特氏杆菌的的最低检出浓度为107 cfu/ml。进行6种混合菌微波共提取时,其中5种病原菌的特异性基因都能扩出,并且条带很亮。该微波提取方法具有快速,简单,高效的特点,能满足大部分食源性病原菌的常规PCR检测需求,大大节约了时间和成本,具有广泛的适用性,为细菌快速分子检测提供了简便手段。     

牛肉及其制品中肉类掺假的荧光PCR鉴别体系优化

根据可能用于牛肉及其制品掺假的肉类原料(猪肉、鸡肉和鸭肉),设计了4对引物和探针.经过测试其具有良好的特异性后,对影响各自聚合酶链式反应(polymerase chain reaction,PCR)扩增的5个影响因素包括Mg2+浓度、Taq DNA聚合酶活、dNTPs浓度、引物和探针浓度以及模板DNA用量等进行了优化,确定了最佳的PCR扩增体系.同时,根据优化后的结果,对特异性进行进一步的测试,并使用建立的优化荧光PCR鉴别体系对混合样品和市售样品进行检测,确定整体检出限为0.1%(质量分数),并证明该鉴别体系的实际应用价值.     

应用PCR技术对DMD做基因诊断的研究

杜氏肌营养不良症，是致死性Ｘ链锁隐性遗传病。     

Real-time PCR的研究进展及其应用

本文综述了real-time PCR的原理、定量方法及其在各领域的应用。     

基于区域时变聚类采样机制的物联网 大数据传输算法

为解决传统物联网大数据传输算法存在的网络链路抖动频繁、传输能力受限以及传输误码率大的不足,提出了一种基于区域时变聚类采样机制的物联网大数据传输算法。首先,根据物联网信号发射中常用的LTE-5G信号特性,采取分信道传输及抽样过滤技术来实现数据传输过程中的正交化,消除节点间因信号频率相似而导致的干涉现象,提高网络数据传输效率;随后,几何聚类机制与能量-阈值映射裁决方法,构建了稳定聚类采样方法,利用功率最佳及能量最佳的方式来实现传输节点的筛选,强化传输链路的稳定性能,降低因链路抖动而导致误码的风险。仿真实验结果表明:与常见的超高斯频率漂移传输提升机制(Transmission Mechanism of Super Gauss Frequency Drift Transmission, SGFD-T机制)及拉普拉斯信道滤波传输机制Laplasse Channel Filtering Transmission Mechanism,LCF-T机制)相比,所提算法具有更大的上传带宽和更低的数据采集错误率与误码率。     

MGB probe assay for rapid detection of mtDNAl1778 mutation in the Chinese LHON patients by real-time PCR

Objective： Leber＇s hereditary optic neuropathy （LHON） is a maternally inherited degeneration of the optic nerve caused by point mutations of mitochondrial DNA （mtDNA）. Many unsolved questions regarding the penetrance and pathophysiological mechanism of LHON demand efficient and reliable mutation testing. This study aims to develop a minor groove binder （MGB） probe assay for rapid detection of mtDNA11778 mutation and heteroplasmy in Chinese LHON patients by real-time polymerase chain reaction （PCR）. Methods： Forty-eight patients suspected of having LHON and their maternal relatives underwent a molecular genetic evaluation, with 20 normal individuals as a control group at the same time. A real-time PCR involving two MGB probes was used to detect the mtDNA 1 1778 mutation and heteroplasmy. A linear standard curve was obtained by pUCmLHONG and pUCmLHONA clones. Results： All 48 LHON patients and their maternal relatives were positive for rntDNA11778 mutation in our assay, 27 heteroplasmic and 21 homoplasmic. Eighteen cases did not show an occurrence of the disease, while 9 developed the disease among the 27 heteroplasmic mutation cases. Eleven did not show an occurrence of the disease, while 10 cases developed the disease among 21 homoplasmic mutation cases. There was a significant difference in the incidence between the heteroplasmic and the homoplasmic mutation types. The time needed for running a real-time PCR assay was only 80 min. Conclusion： This real-time PCR assay is a rapid, reliable method for mtDNA mutation detection as well as heteroplasmy quantification. Detecting this ratio is very important for predicting phenotypic expression of unaffected carriers.     

Detection of genetically modified ingredients in seed samples from Heilongjiang soybean field

A total of 210 soybean samples collected from different areas of Heilongjiang Province were analyzed by PCR to detect the presence of RR (Roundup Ready) soybean ingredient. Results showed that CaMV35S promoter was not detected in all samples. CP4-EPSPS gene was amplified in 13 samples, but all of them were proved to be false positives in restriction endonuclease digestion analysis of PCR products. Further analysis by nested PCR indicated that there were no RR soybean ingredients in the samples. Besides, the amplified fragments of 0xl by CP4-EPSPS primers were sequenced, and the results confirmed again the fact that this fragment was not from CP4-EPSPS gene.     

传送流的软实时复用

MPEG2是由运动图像专家组织提出的图像及其伴音的通用编码标准,该标准在广电网络获得大量应用,MPEG2标准的研究是本论文的基础,论文对与本课题相关的MPEG2标准进行了说明,给出了软件实现传送流实时复用的方法,并详细介绍了其协议处理和时钟处理的原理和实现.最后对算法的性能和可行性进行了分析,得出实验结论.     

雏鹅新型病毒性肠炎病原快速检测技术的建立及应用

根据小鹅瘟病毒(GPV)和鹅源禽腺病毒(FAV)基因组序列特征分别设计了各自的PCR引物,经对反应条件的优化建立了能同时检测两种病毒的二重PCR方法．该法能从雏鹅新型病毒性肠炎和小鹅瘟疫苗株以及分离的鹅源腺病毒蚀斑克隆株分别扩增出相应的特异性产物,而对其它常见鹅病的病原及正常鹅胚肝组织的扩增结果均为阴性;对来自广西不同地方的36份临床病例样品的检测发现,其中具有雏鹅新型病毒性肠炎典型病变的22份样品均可检出GPV和FAV两种病毒．其它的有4份只检出GPV,3份只检出FAV;12份健康雏鹅肝组织中仅有一份检测到GPV;健康成鹅泄殖腔中GPV和FAV的检出率分别达44．29％、37．14％二者都有的占1．43％．结果表明,雏鹅新型病毒性肠炎的病原可能包括GPV和FAV两种病毒;建立的PCR诊断技术具有快速、敏感、特异的特点,可用于该病征的临床快速鉴别诊断以及流行病学的调查研究．