Nov 07, 2017
Version 2
DNA extraction of Nephila clavipes using the Wizard Genomic DNA Purification kit (Promega) V.2
- 1University of Campinas
Protocol Citation: Luiz Filipe Bartoleti, Luiz Filipe Bartoleti 2017. DNA extraction of Nephila clavipes using the Wizard Genomic DNA Purification kit (Promega) . protocols.io https://dx.doi.org/10.17504/protocols.io.knxcvfn
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
Created: November 07, 2017
Last Modified: March 05, 2018
Protocol Integer ID: 8631
Keywords: dna extraction, spider, kit, araneae, Nephila clavipes
Abstract
Protocol used to extract whole genome DNA from Nephila clavipes (Araneae: Araneidae) specimens.
Attachments
Macerate 1-8 legs (around 50 mg of tissue) using liquid nitrogen.
Add 600 µL of Nuclei Lysis solution and homogenize.
600 µL Nuclei Lysis solution
Take the microtubes to the dry bath at 65°C for one hour.
01:00:00 Dry bath
65 °C
Add 200 µL of Protein Precipitation solution and homogeníze.
200 µL Protein Precipitation solution
Centrifugate the microtubes at 13000 rpm for 10 minutes.
00:10:00 Centrifugation
Transfer the supernatant to a new microtube and add 600 µL of Isopropyl alcohol.
600 µL Isopropyl alcohol
Keep the tubes at -20°C for at least one hour.
-20 °C
01:00:00 Keeping tubes at -20°C
Centrifugate the microtubes at 13000 rpm for 6 minutes at 4°C.
4 °C
00:06:00 Centrifugation
Discard the supernatant and add 600 µL of absolut etanol.
600 µL Etanol
Homogenize gently for one minute.
00:01:00 Homogenizing
Centrifugate the microtubes at 13000 rpm for 3 minutes at 4°C.
4 °C
00:03:00 Centrifugation
Discard the supernatant.
Let the samples dry.
Rehydrate with 50 µL of Tris-EDTA buffer.
50 µL Tris-EDTA buffer