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:2/11/2016)... Feb. 11, 2016  Wellcentive today announced it ... Portland, Oregon -based community care organization (CCO) ... health analytics, quality reporting and care management solutions ... its team of quality managers, analysts and care ... provider groups serving FamilyCare members. ...
(Date:2/11/2016)... BEACH, Florida , February 11, 2016 /PRNewswire/ ... PositiveID Corporation ("PositiveID" or "Company") (OTCQB: PSID), ... diagnostics, announced today that its Thermomedics subsidiary, which ... progress on its growth plan in January 2016, ... products distributors, increasing sequential monthly sales growth, and ...
(Date:2/11/2016)... ... , ... Global Stem Cells Group, a world ... Cells Network (GSCN) to distribute exosome injection and other biological products to stem ... Dominican Republic, Colombia, Argentina, Nicaragua, Panama, El Salvador, Venezuela, Peru, Ecuador, Paraguay, Puerto ...
(Date:2/10/2016)... Early-career researchers from Indonesia ... Uganda and Yemen ... Indonesia , Nepal , ... Yemen are being honored for their accomplishments in nutrition, psychiatry, ... mentoring young women scientists who are pursuing careers in agriculture, biology and ...
Breaking Biology Technology:
(Date:1/21/2016)... , January 21, 2016 ... a new market research report "Emotion Detection and Recognition ... Software Tools (Facial Expression, Voice Recognition and Others), ... Global forecast to 2020", published by MarketsandMarkets, the ... to reach USD 22.65 Billion by 2020, at ...
(Date:1/18/2016)... Calif. , Jan. 18, 2016  Extenua ... software that simplifies the use and access of ... and go-to-market partnership with American Cyber.  ... brings extensive experience leading transformational C4ISR and Cyber ... and integrating the latest proven technology solutions," said ...
(Date:1/11/2016)... , Jan. 11, 2016 Synaptics Incorporated (NASDAQ: ... solutions, today announced that its ClearPad ® TouchView ... products won two separate categories in the 8 th ... and Best Technology Breakthrough. The Synaptics ® TDDI ... simplified supply chain, thinner devices, brighter displays and borderless ...
Breaking Biology News(10 mins):