Navigation Links
Total RNA Isolation: 100 l Viral Sample From Cultured-Cell Supernatant Containing Virus


AquaPure RNA Isolation Kit
Catalog #732-6370
Yield: Dependent on sample and RNA expression levels


Method
Sample Collection and Handling
Samples may be fresh or frozen. Collect fresh virus and isolate RNA as quickly as possible; store at 18C for not more than 24 hr. For long-term storage, store at -70 to -80C.

Cell Lysis
1. Add 500 l cell lysis solution to a 1.5 ml microfuge tube that contains 100 l virus from cultured-cell supernatant.

2. Cap the tube and vortex for 5 sec to mix thoroughly.

3. Heat sample to 65C for 5 min to complete lysis. Cool sample to room temperature. (This step is optional when testing for certain viruses such as hepatitis C.)

Protein-DNA Precipitation
1. Add 200 l protein-DNA precipitation solution to the viral cell lysate.

2. Invert capped tube gently 10 times and place tube into an ice bath for 5 min.

3. Centrifuge at 13,00016,000 x g for 3 min. The precipitated proteins and DNA will form a tight pellet.

RNA Precipitation
1. Leaving behind the precipitated protein-DNA pellet, pour the supernatant containing the RNA into a clean 1.5 ml microfuge tube containing 600 l 100% isopropanol (2-propanol). If RNA yield is expected to be low (<1 mg), add glycogen as a carrier to the isopropanol. We recommend adding 1 l of glycogen solution (20 mg/ml) per 600 l isopropanol.

2. Cap the tube and mix the sample by inverting gently 50 times.

3. Centrifuge at 13,00016,000 x g for 3 min; depending on yield, the RNA may or may not be visible as a small, translucent pellet.

4. Pour off supernatant and drai n tube on clean absorbent paper. Add 600 l 70% ethanol and invert the capped tube several times to wash the RNA pellet.

5. Centrifuge at 13,00016,000 x g for 1 min. Carefully pour off the ethanol.

6. Invert and drain the tube on clean absorbent paper and allow to air-dry 1015 min.

RNA Hydration
1. Add 20 l RNA hydration solution (20 l will give a concentration of 50 g/ml if the total yield is 1 g RNA).

2. Allow RNA to rehydrate on ice for at least 30 min. Store purified RNA sample at 70 to 80C until use.

3. Before use, vortex sample vigorously for 5 sec and pulsespin. Pipet sample up and down several times to ensure adequate mixing.



back to top
'"/>

Source:


Page: All 1 2

Related biology technology :

1. New Kit Generates Exceptionally Pure Total RNA for RT-PCR
2. High-Throughput Isolation of Total RNA
3. Isolate and Analyze Total RNA from Cells Harvested by Laser Capture Microdissection
4. Fast Isolation of Total RNA from Small Numbers of Cells
5. Total RNA Isolation Protocol 1,2,3
6. Isolate It All: siRNA miRNA Total RNA Native Protein
7. Isolate Total RNA and Protein From the Same Sample
8. Isolate High Quality Total RNA from LCM Samples Suitable for Microarray and qRT-PCR Analysis
9. Recover High Yields of Total Nucleic Acid from Formalin-fixed, Paraffin-embedded (FFPE) Tissue
10. Total RNA from Whole Blood for Expression Profiling
11. RNAPrep - Total RNA Isolation Purification
Post Your Comments:
*Name:
*Comment:
*Email:


(Date:4/27/2017)... California (PRWEB) , ... April 27, 2017 , ... ... new infographic explaining why mass flow controllers based on capillary thermal mass flow ... industrial gas mass flow control applications. Over 80% of all industrial processes—such ...
(Date:4/27/2017)... ... 27, 2017 , ... The Council for Agricultural Science and ... Lusk, a consummate communicator who promotes agricultural science and technology in the public ... explains how innovation and growth in agriculture are critical for food security and ...
(Date:4/26/2017)... CALIF. (PRWEB) , ... April 26, 2017 , ... ... Lajollacooks4u, San Diego’s premiere team-building and cooking events company, offers one-of-a-kind gifts, ... , Menus specialize in California cuisine, and guests leave inspired with new ...
(Date:4/26/2017)... CO (PRWEB) , ... April 25, 2017 , ... LABS, ... offerings were added to its extensive test menu: Nucleic Acid Testing (NAT) for ... time, LABS was only able to offer NAT screening for blood donors under an ...
Breaking Biology Technology:
(Date:4/11/2017)... BEACH GARDENS, Fla. , April 11, 2017 ... identity management and secure authentication solutions, today announced ... contract by Intelligence Advanced Research Projects Activity (IARPA) ... for IARPA,s Thor program. "Innovation has ... onset and IARPA,s Thor program will allow us ...
(Date:4/5/2017)...  The Allen Institute for Cell Science today announces ... portal and dynamic digital window into the human cell. ... application of deep learning to create predictive models of ... a growing suite of powerful tools. The Allen Cell ... publicly available resources created and shared by the Allen ...
(Date:3/30/2017)...  On April 6-7, 2017, Sequencing.com will host the ... hackathon at Microsoft,s headquarters in Redmond, Washington ... developing health and wellness apps that provide a unique, ... is the first hackathon for personal genomics and ... in the genomics, tech and health industries are sending ...
Breaking Biology News(10 mins):