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基因芯片实验 – 译学馆
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基因芯片实验

DNA Microarray Methodology

This animation will demonstrate how DNA microarray experiments are performed.
这个动画将演示DNA微阵列实验是如何进行的
DNA microarrays, sometimes called DNA chips,
DNA微阵列 有时又称为基因芯片
reveal the expression of thousands of genes
可以监测成千上万个基因基于固体基质
on a solid surface, such as a microscope slide.
比如显微镜载玻片的表达水平
In this example, we’ll use yeast
此例中 我们用酵母作为模型系统
as a model system to illustrate one use of microarrays.
来阐述微阵列的一种用途
One common use of microarrays is to determine
微阵列的一个常见用途是
which genes are activated and which are repressed
评估比较两个细胞群体时
when two populations of cells are compared.
测定哪些基因被激活 哪些被抑制
Every gene is measured simultaneously.
所有的基因都是同时测量的
As an example,
例如
we’ll compare what happens to yeast genes when cells are grown in aerobic
我们比较细胞在有氧条件和无氧条件下生长时
versus anaerobic conditions.
酵母基因的差异
The cells grow and adjust which genes need
为了生存 细胞生长和调节时
to be activated or repressed in order to survive.
哪些基因被抑制 哪些基因被激活
Now it is time to isolate the mRNA from both populations of cells.
现在该从两个细胞群中分离mRNA了
The cells are spun in a centrifuge.
细胞在离心机中旋转
Now that the cells have gathered in pellets,
细胞沉降富集后
we remove the liquid but not the cells.
我们移除液体 而非细胞
Next, it is time to extract the mRNA from the cells.
接下来 该从细胞中提取mRNA了
When we add the extraction buffer, the mRNA is released into the solution.
当我们加入提取缓冲液时 mRNA就被释放到溶液中
Next, we remove the RNA and place it in a fresh tube.
接下来 我们吸取RNA置于新试管中
Now, let’s make the cDNA from the mRNA.
现在 我们从mRNA中提取cDNA
Here we see three out of many mRNA molecules from each tube of cells.
这里我们可以看到来自每个细胞管的三个mRNA
Each mRNA is converted into red or green colored cDNA.
每个mRNA都被转换成红色或绿色的cDNA
When the colored cDNA is made, the mRNA degrades.
当彩色的cDNA形成 mRNA降解
Then we combine the red and green cDNA,
然后我们将红色和绿色的cDNA结合起来
mixing both colors into a single tube.
将两种颜色的cDNA混合到一管中
At last, it’s time to look at the DNA microarray.
最后 就可以看看DNA微阵列了
In our experiment, a microarray or DNA chip contains about six thousand spots,
在我们的实验中 一个微阵列或者基因芯片中约包含6000个点
each spot is a different yeast coding sequence from a different gene.
每个点都是来自不同基因的不同酵母的编码序列
Let’s choose three spots at random to follow in detail.
我们随机选择三个点来详细讲解
Each spot is made of DNA that can base pair with its complementary cDNA.
每个点都是由DNA组成的 它们可以与互补的cDNA形成碱基对
Here are partial sequences from each of the three spots we are observing.
这是我们观察到的三个点的部分序列
Now let’s incubate the mixed cDNA with the DNA chip.
现在让我们把混合的cDNA和基因芯片一起培养
For the sake of our example,
作为范例
we’ll zoom in and show that some of the labeled cDNA
我们放大来看一下
have bound to DNA in the spots and formed base pairings.
这些标记过的cDNA与点中的DNA结合并形成碱基对
Here we see green and red cDNA bound to this spot.
这里我们可以看到绿色和红色的cDNA结合在这个点上
Only red cDNA is bound to this spot.
而且只有红色的cDNA结合在这个点上
And only green cDNA bound to this other spot.
而只有绿色的cDNA才能与另一个点结合
In a real experiment, you would not see any of this detail;
但在真实的实验中 你看不到这些细节
you would only see the original microarray.
你只能看到原始的微阵列
Now we must wash off the unbound cDNA
现在我们要洗去未结合的cDNA
to see what has bound to the microarray.
来看看是什么与微阵列结合
Let’s detect the bound cDNA so it can be visualized.
我们来检测一下结合的cDNA 就可以把它可视化了
We begin by placing the microscope slide containing the microarray inside a scanner.
首先 我们将包含基因芯片的显微镜载玻片放入扫描仪中
We’ll examine the next phase of the process,
我们来研究这个过程的下一个阶段
keeping our focus on the three spots we’ve been following.
把我们的注意力集中到我们持续关注的这三点
First, a green laser scans the microarray.
首先 绿色激光扫描微阵列
The resulting image is stored on a computer for later analysis.
生成的图像存储在计算机上 以供后续分析使用
Now it’s time for the red laser.
现在用红色激光
This image is also stored on a computer for later analysis.
这幅图像也储存在计算机上 以供以后分析使用
Now we move to the analysis phase.
现在我们进入分析过程
After we eject and safely store the microscope slide,
我们取出并安全储存显微镜载玻片之后
we retrieve the red and green images from the computer
从计算机上检索红色和绿色的图像
and create a merged visualization.
创建合并的可视化图像
In the merged image we see an aerobic gene labeled in green,
在合并后的图像中 我们可以看到有氧基因的则标记为绿色
an anaerobic gene labeled in red,
无氧基因的则标记为红色
and a gene labeled in yellow that was expressed in both aerobic and anaerobic conditions.
而在有氧条件下和无氧条件下都有表达的基因则标记为黄色
This is one example of how DNA microarrays are used.
这是DNA微阵列应用的其中一个案例
In an actual experiment,
在实际实验中
quantitative analysis would be conducted on all six thousand genes.
定量分析则囊括所有的这6000个基因

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视频概述

简单介绍基因芯片(DNA微阵列)实验。

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视频来源

https://www.youtube.com/watch?v=0ATUjAxNf6U

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