Jul 02, 2022

Public workspaceDNA extraction (BOMB)

DOI
dx.doi.org/10.17504/protocols.io.x54v9ywq1g3e/v1
  • 1KMU
Tsu-Chun Hung
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DOI: dx.doi.org/10.17504/protocols.io.x54v9ywq1g3e/v1
Protocol Citation: Tsu-Chun Hung, Yin-Tse Huang 2022. DNA extraction (BOMB). protocols.io https://dx.doi.org/10.17504/protocols.io.x54v9ywq1g3e/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it’s working
Created: May 27, 2022
Last Modified: September 13, 2023
Protocol Integer ID: 63322
Abstract
DNA extraction (BOMB)
Sample Collection
Sample Collection
3m
3m
Add Amount200 µL of 1mm beads to 1.5ml enppendorf tube

30s
AddAmount200 µL of 0.5mm beads to 1.5ml enppendorf tube

30s
Add Amount225 µL of TE buffer to 1.5ml enppendorf tube
Note
TE buffer is in 4°C fridge

30s
Add Amount375 µL of lysis buffer to 1.5ml enppendorf tube
Note
Lysis buffer is in 4°C fridge


30s
Add Amount267 µL of 10M ammonium acetate to 1.5ml enppendorf tube
Note
10M ammonium acetate is in 4°C fridge

Collect Amount10-20 mg of sample to 1.5ml enppendorf tube
1m
Sample crush
Sample crush
4m
4m
Put 1.5ml eppendorf tube in mixmill for sample crush, at this condition: 30 rpm/s, for 4mins Duration00:04:00
4m
Centrifugation
Centrifugation
3m
3m
Put 1.5ml eppendorf tube in centrifuge for centrifugation, at this condition:Centrifigation17.0 x g, 25°C, 00:03:00
3m
DNA purification
DNA purification
37m 30s
37m 30s
Add Amount350 µL of isopropanol to the 1st well of 96 well plate



30s
Add Amount125 µL of magnetic beads (10 mg/ml) to the 1st well of 96 deep well plate

Note
Shake the bottle and pipetting before using magnetic beads


30s
Add Amount200 µL of sample (lysate) from 15ml centrifuge microtube to the 1st well of 96 deep well plate
Note
USUALLY ADD at LAST

30s
Add Amount400 µL of isopropanol to the 2nd well of 96 deep well plate
30s
Add Amount300 µL of 80% enthanol to the 3rd well of 96 deep well plate
30s
Add Amount300 µL of 80% enthanol to the 4th well of 96 deep well plate
30s
Add Amount100 µL of DD water to the 5th well of 96 deep well plate
30s
Put in the automated DNA extraction machine

34m
After the extraction is done, collect Amount100 µL of the eluted sample as the DNA template for downstream experiments