introduce
The experimental data is completed, how to draw it as a histogram?
This article takes real-time quantitative PCR (qPCR) as an example.
qPCR is a real-time detection of the amplification process through fluorescent signals. Due to its high sensitivity and specificity, and the Ct value of the sample is related to the gene copy number, it has been used as an important verification method in scientific research experiments.
Today, the spicy chicken editor will teach you the whole process from data processing to graphic drawing, you must remember it~
I'm about to advertise, and the complained article will be deleted.
software
Software: Excel Student Edition and GraphPad Prism; this article uses GraphPad Prism7
Use the coded data, for example: the relative expression of the X gene in the nematode liver treatment compared with the control (CG), the relative expression is obtained by preprocessing the data, and then plotted with graphpad.
1. From the qPCR data exported from the instrument, find the result from the data. (7500 Instruments)
2. Since the exported data is sorted according to the sequence of the well plate, it is necessary to sort and filter the selected gene column and calculate the data. The control group and the experimental group were replicated separately. It is convenient to calculate, do not calculate on the basis of raw data. Keep the original data. Values that are too different can be eliminated.
For example (15.1, 15.2, 25.2), although there is no basis for 25.2 to be wrong data,, but I just want to remove it. . .
For specific sorting, see point 3 for details.
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3. Calculate the average of 4 (or 3) biological replicates respectively, and calculate according to the formula in the following figure
Step 1: △Ct = (CT of target gene-CT of reference gene),
The second step: △△Ct (△Ct of the experimental group - △Ct of the control group ),
The third step: 2^-△△Ct value (relative expression, negative △△Ct of 2 , can also be calculated using the function POWER),
【其中Ct值的范围是15-35，这个范围似乎每个实验组的要求不一样，主要看自己】。
数据都是瞎编的。
这里你的出来只有一个值，其实就是GAPDH是默认是 1.
4、相对差异倍数处理好后，打开GraphPadPrism，选择Column，填入Excel中整理好的3次生物学重复实验数据，作图。
5、点击左侧Data1(第4点4)，出现如下图形样式，可选择你自己喜欢的。
6、生物学实验要求进行统计学分析，点击上面菜单栏中的Analyze，选择Column analyses中的t tests，选择右侧要比对的2个样本，点击确定，弹出的对话框中就会显示P value和P value summary。
线虫的实验基本上T检验就能解决大部分。
7、画出表示差异的标识。点击左侧Data1返回柱状图页面，点击菜单栏中的Draw和Write的如下位置，在柱状图上画出——和****(T 打字自己输入** )，某些杂志甚至对图片的字体都做出来了要求。
8、由于软件直接给出的图不好看。甚至不如黑白，我们可以手动进行设置；鼠标分别双击误差线，柱子，坐标轴，更改误差线的颜色、宽度；*的颜色、大小设置；XY轴和柱状图的颜色、宽度、长度等。(遵循哪里不懂点哪里？？！！)
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双击图片(柱子？，哪里想改点哪里)
9、最终图形如下(左图)，你也可以改成彩色(右图)。
10、保存；做好柱状图后，点左侧列表的Data1—File—Export—选择格式—1200dpi—点击OK保存。
杂志对格式要求很严格JPG常用。作图的时候dpi最高就对，组合的时候再按杂志要求。
对于颜色的选择……主要还是多对高分文章的积累。
彩图的画某些杂志是要收费发表的……然后黑白最实用，但是总觉得少了什么。
qPCR的值不仅可以做成柱状图，还可以做成点图，柱子图+点图，还可以做成热图。第五点的时候可以选择。当然数据不好啥也白搭。数据好啥图都行~
知乎言：
The histogram is a post-production tool that everyone is very familiar with, but most people may just use it to observe the exposure distribution trend of a photo. In fact, the information that can be mined by the histogram is very large, and many photo details are hidden in the histogram. Learning to read histograms is a very important lesson.
Are you out of school? It's easy!
Come and try it now!
(This article is also explained in detail in the qPCR chapter of the large project)