Shulou(Shulou.com)06/01 Report--

This article is about how to carry out aCGH chip analysis, the editor thinks it is very practical, so I share it with you to learn. I hope you can get something after reading this article.

ACGH chip is a two-color chip, through the ratio of red and green fluorescence, usually called log2 ratio, to reflect the change of DNA copy number between the test sample and the control sample. The analysis of aCGH chip usually includes the following three steps

1. Normalization

Normalization has two functions, the first is to correct system errors, and the second is to make the log2 ratio of different samples can be compared directly. In aCGH chips, system errors include the following

Intensity bias, error of fluorescence signal strength

Spatial bias, spatial error, as shown in the following figure, there are differences in the distribution of log2 ratio values at different positions on the slide, that is, different spot

Plate bias, the spot on the chip belongs to different plate, and the log2 ratio distribution of different plate is also significantly different.

Background bias, background error, some of the measured fluorescence intensity comes from background fluorescence.

The original chip data can be analyzed in the next step only after normalization.

2. Segmentation

The segmentation step is to divide the genome into non-overlapping regions, and the number of copies in each region is the same, as shown in the following figure

The blue dot represents the log2 ratio value, the red line represents the divided area, and the log2 ratio in each region is corrected to a uniform value, such as the mean. Each divided region is called segment, and the point between different regions is called breakpoint.

3. Calling

Calling means to analyze the change of copy number on each segment. According to the log2 value corresponding to the segment fragment, it can only be concluded that there are three cases in which the copy number of the test sample changes relative to the control sample.

Normal, the number of copies has not changed, usually denoted by the number 0

Loss, which reduces the number of copies, usually denoted by the number-1

Gain, an increase in the number of copies, is usually denoted by the number + 1. When it increases a lot, for diploids, it is usually greater than 5, it is called amplification.

There are many softwares to analyze aCGH chips, and the algorithms of segmention vary with normalization of different softwares, but the steps of analysis are all above. In order to facilitate downstream analysis, there is a file format for storing copy number variation, called SEG, which is shown below.

Is a\ t-separated threading text format, the first column is the sample ID, the second to the fourth column describes the corresponding chromosome region of segment, and the fifth column represents the number of probes contained in the segment region. In NGS-based CNV analysis, this column represents the number of bin intervals contained, and the sixth column represents the mean value of log2 ratio in the region.

The above is how to carry out aCGH chip analysis, the editor believes that there are some knowledge points that we may see or use in our daily work. I hope you can learn more from this article. For more details, please follow the industry information channel.

Welcome to subscribe "Shulou Technology Information " to get latest news, interesting things and hot topics in the IT industry, and controls the hottest and latest Internet news, technology news and IT industry trends.