Setting up a DNA extraction lab is quite difficult for the novice. However, it is not a big task yet we have to select all the utilities very carefully. which instruments are required, which chemicals are required and what is the SOP for different types of sample to proceed, we have to set up all the things very accurately.
Suppose you are new to the DNA extraction lab, you just started your experiment and in the middle of the experiment you realize that you do not have ethanol, then what?
and it can happen to newbies.
In this article, we are giving you a complete guide on how to set up a DNA extraction lab. what are the instruments and reagents to be used in the lab and what are the other utilities?
In the very beginning decide your goal.
What type of DNA extraction you have to perform whether you are doing all the samples or just one type of sample.
If you want to do DNA extraction from all the types of biological samples such as blood, plant tissue, buccal smear, bacterial load or forensic samples, probably you need more advanced setup.
Now first understand what is DNA extraction.
“DNA extraction is the process of isolating or extract DNA from the cell or from the other cell organelles.”
DNA extraction is complete in the following steps:
- Lysis of cell wall or cell membrane.
- Lysis of nuclear membrane.
- Extracting the DNA from the other cell debris
- DNA precipitation
- Dissolving DNA.
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These are the general outline of DNA extraction, all the steps must be followed in any types of DNA extraction protocols.
For more detail on the DNA extraction and different types of DNA extraction method read our article: Different types of DNA extraction methods
This article contains all the information on DNA extraction and different types of methods used in DNA extraction.
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let us assume that our DNA extraction lab is for all types of samples.
We need three types of utilities for that:
- Plasticware and glassware
Chemicals for DNA extraction lab:
Detergents and salt are the best choices for cell lysis. CTAB, SDS, NaCl, KCl, Tris and EDTA are some of the common chemicals must require for the DNA extraction.
The list of the chemicals and its role is enlisted into the table below,
|Tris||DNA extraction buffer, DNA elution buffer|
|EDTA||DNA extraction buffer, DNA elution buffer|
|SDS||DNA extraction (cell membrane and nuclear membrane lysis)|
|CTAB||DNA extraction (cell membrane and nuclear membrane lysis)|
|KCl||DNA extraction buffer (separating DNA)|
|NaCl||DNA precipitation and DNA extraction|
|MgCl2||DNA extraction (protects DNA)|
|Nonidet P40||DNA extraction|
|Chloroform||Prevent shearing of DNA|
|Isoamyl alcohol||Separates DNA from other impurities|
|Proteinase K||Protein digestion|
|Liquid nitrogen||Breaking the plant tissue|
|Boric acid||Agarose gel electrophoresis|
|Agarose powder||Agarose gel electrophoresis|
|Acetic acid||Agarose gel electrophoresis|
|EtBr||Agarose gel electrophoresis|
|Bromophenol blue||Agarose gel electrophoresis|
DNA extraction buffer, preparation of phenol
These are the general chemicals required in any DNA extraction lab.
Instruments for DNA extraction lab:
|High-speed centrifuge||For centrifugation of samples|
|UV transilluminator||Result analysis|
|Agarose gel electrophoresis unit||For gel electrophoresis of DNA|
|Nanodrop light spectrophotometer||DNA quantification and purity check|
|Qubit fluorometer||DNA quantification|
|pH meter||For checking pH|
|Weigh balance||Weighing the chemicals|
The figure below is the general setup of the DNA extraction lab.
Outside the DNA extraction lab:
Before the sample enters into the DNA extraction lab, the sample must be labelled and recorded into the laboratory management system.
For that generate the unique code for the laboratory or unique barcode for the lab.
Also, the lab does have the sample collection facilities, in the case when a patient directly visits the lab. for that, we do have a syringe, needle, cotton, blood collection tube and spirit outside the lab.
If the sample is blood, the sample must be collected in the EDTA vial.
The fire safely setup must be present outside the lab.
Also, the lab must be labelled with the essential labels.
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Inside the DNA extraction lab:
Starting with the sample preparation, the sample must be labelled once again for the DNA extraction lab. This place is dedicated to sampling preparation and addition of other chemicals.
After that place centrifuge, water bath and vertex next to the sample preparation area (see the figure).
After that into the next side of the lab, place the weigh balance first followed by the oven, electrophoresis unit and UV transilluminator.
Here at this station, the extracted DNA sample is electrophoresis. The gel is prepared and the DNA is run on the agarose gel electrophoresis, the DNA is analysed under the UV transilluminator.
Next to that is the DNA quantification.
Once the electrophoresis is done, the sample is sent to the quantification. Here the purity and the quantity of the extracted DNA is checked and then it is sent to the next station.
Now our sample is ready for the PCR and other downstream applications. The DNA is sent to the storage (freeze or deep freeze). From the sample, our DNA is extracted, now the DNA can send to the next section of our genetic lab.
The general set of the genetic lab is shown in the figure below,
The DNA extraction lab is an essential part of the genetic lab. Starting with the DNA extraction lab, it opens into the storage room.
The storage room contains freeze, deep freeze and utility racks. once the DNA is extracted from the DNA extraction lab, the DNA is quantified and stored into freeze for short term storage and deep freeze for long term storage.
The storage room opens into the reaction preparation room. The reaction preparation room contains the PCR work station where the PCR reactions are prepared.
The reaction preparation room opens into the template addition room, where the DNA template is added to the PCR reaction and placed into the PCR machine.
The described setup for the extraction lab is as per the NABL guideline for the genetic lab. Here, as per the NABL guideline, the genetic lab essentially required a dedicated template addition room.
Because in the reaction preparation lab and DNA extraction lab, the chance of the cross-contamination is very high.
Note: Post-amplification area is not shown in the figure above.
Glassware and plasticware for DNA extraction lab:
All the other utilities such as glassware and plasticware are as important and the main utilities. 500ml flask, 1000ml flask, 100ml measuring cylinder, 1000ml measuring cylinder and beakers are some of the commonly used glassware.
1ml, 200mcrolter, 10microliter tips and 2ml and 1.5 ml Eppendorf tubes are required in the DNA extraction lab.
|DNA extraction kits (for back up)|
|DNA extraction solutions (ready to use for all type of sample)|
|Ready to use TE buffer (Back up)|
|Ready to use gel loading buffer|
|Blood collection tubes|
|Syringe and needle|
Records and registers:
As per the standard guideline records and registers are an important part of the DNA extraction or genetic lab. Maintain the record of the patients, sample and its proceeding times. Also, create registers such as stock register, sample register, outsource register, utility register, chemical register and other stock registers.
Maintain the register by entering the date of usage, date of stock completion and breakdown date of instruments. Create separate files for all the manuals of the instruments and other documents.
We have covered articles on the preparation of different types of solutions for different types of DNA extraction methods.
Chemical preparation for Phenol chloroform DNA extraction method:
Solution preparation for CTAB DNA extraction method:
TE buffer preparation:
DNA extraction buffer preparation:
Further, for the backup source purchase ready to use DNA extraction kits, DNA extraction solutions, TAE and TBE buffer, TE buffer and other solution so that in case of failure in daily protocol your time and sample does not get wasted.
DNA extraction is one of the crucial steps in the post PCR and PCR analysis. Hence it is very important to set up a DNA extraction lab very preciously. The yield and the quality of the DNA depend on the structure of the DNA extraction lab. Furthermore, it also depends on the SOPs (How you maintain) and the protocols (How you standardise) of the lab.
Remember, always prepare your own lab protocols and SOPs.
Our experts Mr Ravi Parmar is a NABL trainer and Mr Tushar Kachhadiya is a NABL expert, you can contact us for genetic lab set up. For lab set up Contact us here.