hello friends
I want to detect some rare genes in tumoral samples . when I used TAKAR kit for my real time PCR, I could find in some samples but I changed my Kit for real time PCR, it is ABI, and now I cannot detect those genes in my samples. what do you think? I thought ABI kit is more sensitive and because of that I changed kit , but now I am not sure about it. I have to say the cDNA concentration and also primer concentration are same in both kits.
It may be the RT rather than the qPCR that is causing the variability in your results. Different RT enzymes generate cDNA at different efficiencies and they can also be assay dependent. Low abundance targets may not be converted using certain RT enzymes. You need to validate your approach for the particular samples and GOI targets you are interested in and then stick with that approach. Unfortunately there is no one kit that fits all scenarios. Check out the following publication for more information:
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3779174/
I agree with Rebecca's comments.
Nevertheless, if you were happy with your first kit, it would be worth going back to it.
سلام
قدم به قدم جلو برید؛
اول دوباره با همون نمونه ای جواب نمیده با تاکارا تکرار کنید، اگر جواب داد یعنی باید مسترمیکس جدید رو ست کنید، یعنی به غلظت و رقت های مطلوب برسید که فقط راهش تکرار است.
اگر جواب نداد یعنی یا دستگاه ایراد داره با دیگه نمونه کیفیت مطلوبی نداره، بنابرین کیتتون رو با نمونه بقیه دوستان چک کنید یا دستگاه دیگه ای رو امتحان کنید
کلا زنجان خوبیش اینه، استاد میشین میان بیرون!!!
The TAKARA kits have been shown to be more sensitive than most other kits. TAKARA is more expensive however - is that why you switched kits? Different kits have been known to give widely differing sensitivities in general.
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