I have done some Sugarcane Streak Mosaic virus identification tests for sugarcane varieties. The band intensity is differ from variety to variety. How can I measure the band intensity? A software is in the package. Gel images taken by Vilber gel doc system.
Dear Ms Perera,
The bands' intensity is not so important for measurements of virus density. The bands mean there are viruses in your samples. The intensive bands can only give idea about your target DNA fragment copied well in RT-PCR. Weak bands mean that your NA is not copied well in PCR or it could not been isolated well.
Good studies and wishes
Hi Malika, I would prepare at least 3 dilutions of a known concentration of DNA fragment (in duplicate) and run them concurrently with your PCR product on the gel. You can measure the intensity of each band using software that either comes with the gel imaging system or ImageJ as mentioned by Md Abdur Rahim . Then you plot the band intensity on y-axis vs concentration on x-axis to obtain the calibration curve. Once done you can apply a linear trendline and obtain the equation (y=mx+n) and use it to convert the band intensity of your sample to a concentration. Band intensity of sample should be within the range used for the calibration curve. If it is not you may have to dilute the sample or use higher DNA concentration for your standard.
Hi Malika Perera
have you been able to develop / follow a protocol to measure the band intensity?
Kind regards, Rob.
Sorry, unable to develop that protocol Prof. Robert. At that time my under graduate student discard all the cDNA samples. Disease identification level is enough for her study, according to the supervisor. So I stopped the project up to that level. Try to start it regarding 2019 research program.
In addition, supplier of gel doc system do not have knowledge on the software. I try to find it in the web. But still couldn't. No any supportive doc with instrument. I can't operate the software.
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