端粒和端粒酶在人类干细胞和癌症中的作用 - Elizabeth Blackburn P2
本视频由科普中国和生物医学大讲堂出品
Elizabeth Blackburn (UCSF) Part 2: Telomeres and Telomerase in Human Stem Cells and in Cancer
Telomerase, a specialized ribonucleprotein reverse transcriptase, is important for long-term eukaryotic cell proliferation and genomic stability, because it replenishes the DNA at telomeres. Thus depending on cell type telomerase partially or completely (depending on cell type) counteracts the progressive shortening of telomeres that otherwise occurs. Telomerase is highly active in many human malignancies, and a potential target for anti-cancer approaches. Furthermore, recent collaborative studies have shown the relationship between accelerated telomere shortening and life stress and that low telomerase levels are associated with six prominent risk factors for cardiovascular disease.
限制性内切酶消化
限制性酶或内切酶会识别并切开DNA上的某个特定序列。这些酶在细菌中天然存在，用来抵御感染细菌的病毒-噬菌体的攻击。细菌中的限制性内切酶会切开入侵噬菌体的DNA，而细菌本身的DNA由于有甲基化，将不会受到影响。
本短片讲述了限制性内切酶的基本原理：内切酶是如何命名的，已有的酶切位点和突出端类型。并将讲解常用的限制性内切酶消化的逐步操作过程，包括所需组分，添加各组分到混合物中的顺序，以及常用的消化温度和时间。还将提及灭活限制性内切酶来抑制非特异性活性的重要性。短片还给出了操作多酶消化及酶切中使用对照的小技巧。
酶联免疫吸附测定法
酶联免疫吸附检测(ELISA)通常是检测实验样品中是否存在某目的蛋白，以及该目的蛋白的量。目的蛋白的识别归功于抗体，因此ELISA是一个免疫检测。这些抗体通常偶联了一种酶，经过一系列的孵育，洗脱步骤，它们将检测到包被在多孔板孔底部的蛋白。当底物存在时，偶联在抗体上的酶将造成颜色改变，这说明样品中存在目的蛋白。
本短片将讲解ELISA的工作原理，其中讨论了一抗与二抗结合，以及封闭步骤的重要性。在接下来的练习中，视频演示了其逐步操作过程。最后介绍了标准ELISA的几种变化形式，如三明治ELISA和竞争ELISA，以及该方法的实际应用，例如非处方的测孕试纸
PCR:聚合酶链式反应
聚合酶链式反应，或称PCR，是一种通过热循环反应来扩增DNA的技术，热循环是指在固定的时间间隔内温度变化的循环。采用热稳定的DNA聚合酶，PCR可以用DNA的合成原料，脱氧核苷三磷酸合成大量的DNA拷贝。PCR反应包括三步，变性，退火，和延伸。变性是PCR循环的第一步，它将DNA碱基之间的氢键打开来熔解DNA双链而产生单链DNA。退火则是将温度降到足够低，使寡聚核苷酸引物能够结合到DNA模板上。在延伸过程中，DNA聚合酶将合成新的双链DNA。
本短片将介绍PCR的过程。除了逐步讲述如何准备一个常用的PCR反应，还将描述PCR的基本原理。视频展示了一个PCR反应的所需组分，以及引物设计的规则，并给出了保证PCR反应成功的一些有用技巧。
核酸性酶的发现 - 托马斯·罗伯特·切赫
本视频由科普中国和生物医学大讲堂出品
托马斯·罗伯特·切赫，在本次讲座中讲述了他那令人激动的实验，如何发现RNA可以催化来切割和连接生成化学键。在此之前发现，大多数科学家认为，只有蛋白质可以进行催化的生物反应。这项工作的重要性已被公认的1989诺贝尔化学奖。
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蛋白激酶的结构 - Susan Taylor
In this lecture, I have given an overview of protein kinase structure and function using cyclic AMP dependent kinase (PKA) as a prototype for this enzyme superfamily. I have demonstrated what we have learned from the overall structural kinome which allows us to compare many protein kinases and also to appreciate how the highly regulated eukaryotic protein kinase has evolved. By comparing many protein kinase structures, we are beginning to elucidate general rules of architecture. In addition, I have attempted to illustrate how PKA is regulated by cAMP and how it is localized to specific macromolecular complexes through scaffold proteins.
蛋白激酶的调控与定位- Susan Taylor
In this lecture, I have given an overview of protein kinase structure and function using cyclic AMP dependent kinase (PKA) as a prototype for this enzyme superfamily. I have demonstrated what we have learned from the overall structural kinome which allows us to compare many protein kinases and also to appreciate how the highly regulated eukaryotic protein kinase has evolved. By comparing many protein kinase structures, we are beginning to elucidate general rules of architecture. In addition, I have attempted to illustrate how PKA is regulated by cAMP and how it is localized to specific macromolecular complexes through scaffold proteins.
GTP酶反应和疾病
In the second part of Dr. Wittinghofer's talk he explains the link between GTPases and disease. Ras is both a key molecule in regulating normal cell growth and an oncogene in unregulated cancer cell growth. Mutations in Ras that prevent the hydrolysis of GTP to GDP lock Ras into an active state rendering it independent of upstream growth factor signals. Biophysical studies from Wittinghofer's lab solved the multiple steps in the hydrolysis of GTP to GDP and explained why particular mutations in either Ras or Ras-GAPs cause unregulated activation of Ras and tumor formation. Examples of other G-proteins that are unable to hydrolyse GTP and result in different diseases such as Retinitis Pigmentosa, are also presented.
聚合酶链反应的简介
What is Polymerase Chain Reaction (PCR)? PCR is a molecular technology developed by Nobel laureate Kary Mullis in the 1980s that allows the fast and inexpensive amplification of DNA fragments in vitro. It has become a fundamental tool in genetic and molecular research.
DNA聚合酶链式反应
What are DNA polymerases? DNA polymerases are enzymes responsible for assembling nucleotides to create new DNA molecules. These DNA polymerases are indispensable to cell divisions as they duplicate the genetic information that would be passed to the next generation of daughter cells. Throughout the years, scientists have utilized different variants of DNA polymerases for different applications.