Apr 27, 2022
Genomic RNA extraction by TRIzol method by using Drosophila melanogaster gut
- 1Kuvempu University
Protocol Citation: Mohamed Sabeelil Islam 2022. Genomic RNA extraction by TRIzol method by using Drosophila melanogaster gut. protocols.io https://dx.doi.org/10.17504/protocols.io.dm6gpb92plzp/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
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Created: April 04, 2022
Last Modified: April 28, 2022
Protocol Integer ID: 60261
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Abstract
Simple method followed for RNA extraction of Drosophila melanogaster larval(3rd instar) gut for CDNA
Guidelines
ware gloves while extraction
Dissection was used to isolate the larval gut of Drosophila 3rd Instar.
(around 10-15 gut was taken)
The dissected gut was taken in a 1.5ml ependorf tube which contained 200μL of TRIzol.
(For a total of 800μL or 0.8 mL of TRIzol needed for the extraction)
To avoid spurt, 200μL were taken.
Homogenize by using a micropistle
Add 600μL or 0.6mL of TRIzol followed up by adding 200μL or 0.2mL of Chloroform
(after adding vortex it) and keep it for 5 minutes incubation
Centrifuge at 12000x for 10minutes at 4ºC
Take supernatant into new tube and add equal amount of isopropanol
Centrifuge at 12000x for 10minutes at 4ºC
discard the supernatant and retain the pellet
wash the pellet by using 1ml of 75% cold ethanol (twice)
(after adding ethanol vortex the solution and centrifuge at 7500x for 5 minutes)
Retain the pellet, add RNase free water and check the purity