基于微流控芯片的核酸等温扩增技术研究进展

核酸等温扩增技术是一种在恒温体系内对核酸进行高效扩增的分子扩增技术,它能够在短时间内实现目的基因的指数增长.微流控芯片(microfluidic chip)技术是把研究样品制备、核酸富集、纯化和检测等多个操作步骤集成到一块“微型化”的芯片上,经自动化处理,得出实验结果,即“样品进,结果出”.将核酸等温扩增技术与微流控芯...     

用于新型冠状病毒检测的核酸等温扩增技术研究进展

核酸检测作为新型冠状病毒肺炎(COVID-19)筛查诊断和病情监测的主要手段,在疫情防控中发挥了重要作用。虽然实时荧光定量PCR被认为是新型冠状病毒(SARS-CoV-2)核酸检测的金标准,但其依赖荧光定量PCR仪且扩增检测时间较长,难以实现现场快速检测。因此许多基于核酸等温扩增的SARS-CoV-2检测方法相继诞生。等温扩增对仪器温控要求不高,通过与微流控芯片和可视化检测技术结合,可进一步简化操作、降低成本,为SARS-CoV-2现场快速筛查提供有力的技术支撑。本文围绕已报道的SARS-CoV-2等温扩增检测方法原理、检测性能及优缺点进行探讨,为进一步发展SARS-CoV-2现场快速检测平台提供参考。     

基于核酸等温扩增的病原微生物微流控检测技术

病原微生物的快速检测对疫情的预防控制至关重要。基于PCR的病原微生物核酸检测方法克服了传统病原微生物培养方法耗时长、免疫学检测存在窗口期等问题,已成为目前最主要的病原微生物筛查方法。然而,对精确控温热循环仪的依赖却严重限制了其在资源匮乏地区的应用。虽然基于核酸等温扩增的病原微生物检测方法可摆脱对高精度温控设备的依赖,但仍需要进行样本核酸分离提取、扩增与检测等步骤。近年来,微流控技术与核酸等温扩增技术相结合,诞生了多种病原微生物等温扩增微流控检测技术。该技术通过设计芯片结构、优化进样模式及检测方式,实现了病原微生物核酸提取、扩增与检测一体化,并具备多重检测、定量检测等功能,具有对仪器依赖度小、对操作人员要求不高、样本量需求小和自动化程度高等优点,适合于在多种环境下的病原微生物快速检测。本文从核酸等温扩增原理、进样方式、检测方式等方面对核酸等温扩增病原微生物微流控检测技术进行了综述,以期为病原微生物的快速筛查提供更多的方案思路,提升公共卫生领域对传染性疾病的防控能力。     

综述与专论: 基于核酸适配体传感器的即时检测（POCT）技术

即时检测（point-of-care testing，POCT）是一种检测成本低、检测速度快、准确度高、能自我采样获得临床诊断结果的新型诊断技术。该技术在临床诊断、病情监控与疫情防控等领域发挥了重要作用。核酸适配体是一种能够特异性识别多种靶标的分子探针，具有易合成、批间差异小、易实现信号放大等突出优势，是生物医学传感器中重要的分子识别元件。本文概述了核酸适配体探针的现有筛选方法和进展，总结了核酸适配体POCT传感器信号放大策略，着重介绍了各类核酸适配体传感器在POCT领域的应用现状，并对核酸适配体POCT传感器的发展前景进行了展望。     

基于环介导等温扩增技术的微流控芯片在病原菌检测方面的研究进展

环介导等温扩增(LAMP)技术是一种新兴的核酸恒温扩增技术,与微流控芯片技术相结合,可实现对病原菌的快速检测,具有特异性强、灵敏度高、操作简单等优点。本文根据不同终产物的检测方法对目前检测病原菌的相关微流控LAMP芯片进行了分类与介绍,并对技术的改进和存在的问题进行了分析,以期为后续的相关研究提供参考。     

等温扩增技术及其结合CRISPR在微生物快速检测中的研究进展

等温扩增技术因其对仪器依赖性低、核酸扩增高效等优势,非常适合于快速检测,已在微生物快速检测领域得到了广泛应用。本文从核酸提取、等温扩增(以环介导等温扩增技术(Loop-mediated isothermal amplification,LAMP)和重组酶聚合酶扩增技术(Recombinase polymerase amplification,RPA)为例)和产物检测角度,就近年来核酸等温扩增技术的发展及其在病原微生物核酸快速检测领域的应用进行综述,并概述了核酸等温扩增技术与CRISPR(Clustered regularly interspaced short palindromic repeats)基因编辑技术相结合的最新研究成果,为这些新兴技术的研究和未来的发展提供新思路。     

分子即时检测(POCT)技术及其在新发传染病中的应用*

分子即时检测(point-of-care testing,POCT)技术具有灵敏度高、分析速度快、体积小、检测成本低廉等特点,在分子诊断领域受到广泛关注。近年来,分子POCT技术的发展与应用在应对新发、突发传染病,保护人类生命健康方面具有重大意义。介绍近五年来新兴的分子POCT技术,总结新兴分子POCT技术的最新研究进展及应用前景,分析POCT技术的优势与面临的挑战,探讨提高其检测灵敏度和选择性的技术策略。     

连续流动式PCR微流控装置的研究

介绍了基于薄膜加热器的新型连续流动式PCR微流控装置的设计与制作;讨论了退火温度、PCR反应试剂(引物、Mg2 、dNTPs以及Taq DNA聚合酶)浓度以及PCR溶液的流动速度等对连续流动式PCR反应的影响;结果发现反应试剂影响连续流动式微流控PCR扩增的行为不同于它们影响传统PCR的行为,在较宽的浓度范围内都不会引起非特异性扩增。除此之外,在15 min内能成功对249 bp的人类β-肌动蛋白基因进行扩增,扩增速度比传统PCR快;通过低热容量的薄膜加热器来维持三个温度区带的恒温,完成33个循环的连续流动式PCR扩增能量消耗小于0.0088 kW.h,比传统PCR仪低得多,新研制的PCR微流控装置有可能成为便携式装置。     

超快脉冲控制PCR（upPCR）技术及应用

在精准医疗、个性化医疗的大背景下,分子诊断在病原体检测、肿瘤诊断、优生优育、环境保护、食品安全等领域的应用越来越广泛,并逐渐向操作简单、快速准确、低成本、适用于基层及家庭使用的分子即时检测（point-of-care testing,POCT）方向发展。超快脉冲控制PCR（ultra-fast pulse-controlled PCR,upPCR）是实时荧光定量PCR（qPCR）技术的延伸和升级,该技术利用能量脉冲控制扩增反应中的金属加热元件（主要是纳米金）,在几百微秒内完成溶液局部微环境的快速升温,实现模板DNA的解链变性,停止加热后反应微环境可被周围溶液快速冷却到聚合酶的延伸温度,实现引物退火和模板DNA的扩增,单个变性-扩增循环仅有1.5~5 s,远快于传统PCR（约90 s/循环）,从而能够极大地加快扩增反应速度。upPCR技术在保留了传统qPCR高灵敏度、高特异性和多重检测等优势的基础上,增加了超快速（低于15 min）、设备简单等新优势,非常适合用于基层检测等分子POCT场景。本文主要对upPCR技术的原理、设备、核心原料及在分子诊断中的应用进行综述,并对该技术存在的优缺点,以及未来的技术发展和应用趋势进行了讨论。     

食源性病毒核酸恒温检测技术研究进展

食源性病毒已成为全球引发食品安全事件的重要病原,对新型检测技术的不断发展提出了严峻的挑战.早期PCR技术在病原检测领域中的应用,推动了对食源性病毒的全面认识.近年来核酸恒温检测技术发展迅速,包括环介导等温扩增技术、重组酶聚合酶扩增技术、核酸序列依赖性扩增技术、链置换扩增技术、滚环扩增技术等,在抗复杂基质干扰、装备要求低...     

Recent advances in the study of Kaposi's sarcoma-associated herpesvirus replication and pathogenesis

It has now been over twenty years since a novel herpesviral genome was identified in Kaposi's sarcoma biopsies. Since then, the cumulative research effort by molecular biologists, virologists, clinicians, and epidemiologists alike has led to the extensive characterization of this tumor virus, Kaposi's sarcoma-associated herpesvirus(KSHV; also known as human herpesvirus 8(HHV-8)), and its associated diseases. Here we review the current knowledge of KSHV biology and pathogenesis, with a particular emphasis on new and exciting advances in the field of epigenetics. We also discuss the development and practicality of various cell culture and animal model systems to study KSHV replication and pathogenesis.     

The structure, reproduction and taxonomy of Vidalia and Osmundaria (Rhodophyta, Rhodomelaceae)

Comprises species occurring mostly in subtidal habitats in tropical, subtropical and warm-temperate areas of the world. An analysis of the type species, V. spiralis (Sonder) Lamouroux ex J. Agardh, a species from Australia, establishes basic characters for distinguishing species in the genus. These characters are (1) branching patterns of thalli, (2) flat blades that may be spiralled on their axis, (3) width of the blade, (4) primary or secondary derivation of sterile and fertile branchlets and (5) position of sterile and fertile branchlets on the thalli. Application of the latter two characters provides an important basic method for separation of species into three major groups. Osmundaria , a genus known only in southern Australia, was studied in relation to Vidalia , and its separation from the Vidalia assemblage is not accepted. Species of Vidalia therefore are transferred to the older genus name, Osmundaria. Two new species, Osmundaria papenfussii and Osmundaria oliveae are described from Natal. Confusion in the usage of the epithet, Vidalia fimbriala Brown ex Turner has been clarified, and Vidalia gregaria Falkenberg, described as an epiphyte on Osmundaria pro/ifera Lamouroux, is revealed to be young branches of the host, Osmundaria prolifera.     

New or rare chromosome counts in Artemisia L. (Asteraceae, Anthemideae) and related genera from Kazakhstan

Fifteen chromosome counts of six Artemisia taxa and one species of each of the genera Brachanthemum, Hippolytia, Kaschgaria, Lepidolopsis and Turaniphytum are reported from Kazakhstan. Three of them are new reports, two are not consistent with previous counts and the remainder are confirmations of very scarce (one to four) earlier records. All the populations studied have the same basic chromosome number, x = 9, with ploidy levels ranging from 2x to 6x. Some correlations between ploidy level, morphological characters and distribution are noted.     

肝癌中HBV和HCV基因和抗原的分布及意义

采用原位分子杂交方法检测HCV RNA及HBV X基因;采用免疫组织化学方法研究HCV核心抗原,非结构区C33c抗原及HBxAg在肝细胞肝癌中的定位及分布.结果表明(1)HCV RNA、HBV X基因在肝细胞肝癌组织检出率分别为40%(55/136)和82%(112/136).HCV RNA定位于癌细胞的胞浆内,阳性细胞呈散在、灶状及弥漫分布三种形式;HBV X基因在肝癌细胞中的分布呈胞浆型、核型及核浆型,阳性细胞也呈上述三种分布形式;(2)HCV C33c抗原、核心抗原在肝细胞肝癌中的阳性率为81%(133/164)及86%(141/164).C33c抗原定位于癌细胞及肝细胞的胞浆内;核心抗原既定位于癌细胞核中,又可定位于胞浆中.C33c抗原阳性细胞以灶状分布为主;而核心抗原阳性细     

Selection with two alleles and complete dominance

For a plant selection model with frequency-independent viabilities, fertilities and selfing rates, it is shown that apart from global fixation, for certain parameter combinations a protected polymorphism and facultative fixation (either allele may become fixed according to initial frequencies) may both occur. Facultative fixation requires different selling rates for the dominant and recessive type. Protection of the polymorphism requires resource allocation for male and female function. In this connection the problem of purely genetically caused population extinction is discussed.
For general frequency dependence and regular segregation, the chances for establishment of a completely recessive gene are compared to those of a completely dominant gene. It is proven that the process of establishment of the recessive gene, despite a fitness advantage, may be considerably endangered by drift effects if random mating prevails. The recessive gene may reach the same effectivity in establishment as a dominant gene, only if the recessive homozygote mates exclusively with its own type during the period of establishment.