• The smallest PCR reaction volume that I know is provided by PCR analyzer AriaDNA (1.0- 3.6 μl) https://www.lumexinstruments.com/catalog/pcr_analysis/aria_dna.php

Ahmed H. Shatta There are optimization procedures for finding the lowest volume which efficiently amplifies your DNA, which can be based on orthogonal experimentation. I would be happy to give you an elaborate explanation in the messages if required.

Explanation - Efficiency of PCR reaction can be better and can be worked with reduced volume also if your proportions of input DNA and primer/probe concentrations are proportionate even in low volumes so that the plateau phase of PCR doesn't occur before the average cycle count you are providing.

Suggestion based on explanation - So you can provide reduced primer/probe concentration and also reduce input DNA proportionally so that at least a limited amount of volume can be reduced.

what is you volume reaction?

Dear

Takwa Wannassi

Sorry for the late reply. My reaction's volume is 20 microliters ( 18 μl for master mix & 2 μl DNA )

BR.

For example- you can do a 10ul or 40ul instead of a 20ul but you will have to use 1/2 or double volume of all your PCR components and reagents. I would recommend however to increase your DNA volume. By this you can scale up or scale down your pcr reaction volume from the established protocol.

Ahmed H. Shatta We used small volume (5-10 μl) for PCR under mineral oil.

Aravind Kumar N

Dear,

I am doing a PCR with the NEBNext Ultra DNA library Prep kit for Illumina. However The cDNA that I want to amplify is attached to beads and thus tend to sediment. To avoid this, I divise my cDNA on beads in 4 and run 4 PCR for one sample. I would like to reduce the volume of my reaction.

I currently mix :

24µL of cDNA on Beads

+ 25µL of master mix

+ 0.5µL of each primers (that I reduced to eliminate the excess remaining primers after PCR)

for a total reaction of 50µL.

Any advice please ? Thanks,