Hello Reham Karam,

Follow the steps below for extracting best quality RNA  from stool.

1. Suspend 200 mg of stool in 600 ul of 0.9% NaCl containing 20 uMolar beta mercapto-ethanol. ( This is the maximum recommended 

2. Mix it by pipetting gently, centrifuge at 10000 rpm for 5 minutes.

3. Collect the supernatant and discard the pellet.

4. Filter the solution in 0.22 um syringe filter. 

Follow the QIAGEN protocol next https://www.qiagen.com/in/resources/resourcedetail?id=15aa2b1f-6047-4e9d-9d68-59ea58e248dd&lang=en

Hope it will help you

Biswaranjan

Hello Reham Karam,

Follow the steps below for extracting best quality RNA  from stool.

1. Suspend 200 mg of stool in 600 ul of 0.9% NaCl containing 20 uMolar beta mercapto-ethanol. ( This is the maximum recommended 

2. Mix it by pipetting gently, centrifuge at 10000 rpm for 5 minutes.

3. Collect the supernatant and discard the pellet.

4. Filter the solution in 0.22 um syringe filter. 

Follow the QIAGEN protocol next https://www.qiagen.com/in/resources/resourcedetail?id=15aa2b1f-6047-4e9d-9d68-59ea58e248dd&lang=en

Hope it will help you

Biswaranjan

thank you  for your valuable help

will i need any extra steps before cDNA synthesis

i read about the so called hydroxyapatite

will i help ??

If you are not going for sequencing (NGS) of the total RNA , hydroxyapatite method of rRNA degradation and cDNA normalization is not needed.

thanks alot 

i aim for a sucessful PCR to head forward to  seq so i should add hydroxy apatite