Usually taqman qPCR gene expression assays using taqman fast advanced mastermix require 20ul reaction volumes. I wanted to scale down reaction volumes to 10 or 5ul to save on reagents but I was wondering if this is possible without sacrificing data quality.
Yes you can do with 10ul to save reagents. Reduce everything to half than for 20ul
Make reaction mixture and use 10ul per well for each gene in triplicates or quatruplicates as you do. I will suggest not to go for 5ul per well.
Hello Eli Lawrence
Yes, it is possible.
You can scale down to less than 10ul reactions without compromising the analytical outputs provided you take certain precautions. Since you will be dealing with small volumes you need to use mineral oil to cover the reaction mix to prevent evaporation. Use of well calibrated pipettes becomes important since you will be dealing with small volumes. Use of good quality plastic consumable is essential. Also, there is a probability of inhibitory effects that could arise in the reaction mix which you need to deal with.
I hope this is helpful.
Good Luck.
Yes, you can make 10 ul reaction mixture for taqman! However, during running PCR you need to make sure that you have changed the the total volume to 10ul instead of 20ul.
Best of luck
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