Introduction
Hipure viral RNA kit is suitable for purifying viral RNA from samples such as cell-free body fluid or culture medium. The kit is based on silica gel column purification technology. It requires no toxic phenol chloroform extraction and time-consuming alcohol precipitation in the extraction. The whole extraction process takes only 25 minutes. The kit is suitable for extracting viral RNA from 1-140µl cell-free fluid samples such as serum, plasma, cell-free body fluid or culture medium. The product has successfully extracted hepatitis B A/C, hepatitis C RNA, SARS and HIV. The obtained RNA can be directly used for RT-PCR, Northern hybridization and virus detection.
Details
Specifications
| Features | Specifications |
| Main Functions | Extract viral RNA from 140μl cell-free samples |
| Applications | RT-PCR, Northern hybridization, and various virus detection |
| Purification method | Mini spin column |
| Purification technology | Silica technology |
| Process method | Manual (centrifugation or vacuum) |
| Sample type | Cell-free body fluid or culture medium |
| Sample amount | 140μl |
| Elution volume | ≥15μ |
| Time per run | ≤25 minutes |
| Liquid carrying volume per column | 800μl |
| Binding yield of column | 100μg |
Principle
Hipure silica gel column is based on glass fiber filter membrane with high binding force. Under the condition of high concentration of ionizing agent (such as guanidine hydrochloride or guanidine isothiocyanate), the filter membrane can adsorb nucleic acid through hydrogen bond and electrostatic, while protein and other impurities are not adsorbed and removed. The filter membrane adsorbed with nucleic acid is washed to remove the residual protein and salt. Finally, the nucleic acid adsorbed on the filter membrane can be washed out with low salt buffer (such as buffer TE) or water. The obtained nucleic acid has high purity and can be directly used in various downstream experiments.
The sample is homogenized and lysed in the lysate, and RNA is released into the lysate. The high concentration of guanidine isothiocyanate contained in the lysate denatured and inactivated endogenous or exogenous RNase, while RNA is protected from degradation. The lysate is centrifuged to remove insoluble impurities. After adding ethanol to adjust the binding conditions, it is transferred to the column for filtration. RNA is adsorbed on the membrane of the column, while protein is removed without adsorption. The column is washed with buffer VHB to remove protein and other impurities, washed with buffer RW2 to remove salt. Finally the RNA is eluted by RNase free water. The eluted RNA can be directly used in RT-PCR, Northern blot, poly-A purification, in vitro translation, etc.
Advantages
- Fast – several samples can be extracted in 20 minutes by column method
- High quality – high purity total RNA can be directly used in various sensitive downstream applications
- Safe – no phenol chloroform extraction required
- Sensitive – RNA can be recovered at the level of PG
- High yield – carrier RNA contained in the product maximize the recovery of trace nucleic acid
Kit Contents
| Contents | R417102 | R417103 |
| Purification Times | 50 Preps | 250 Preps |
| HiPure Viral Micro Columns | 50 | 250 |
| 2ml Collection Tubes | 100 | 500 |
| Buffer VRL | 50 ml | 200 ml |
| Carrier RNA | 310 µg | 3 x 310 µg |
| Buffer VHB* | 13 ml | 110 ml |
| Buffer RW2* | 20 ml | 2 x 50 ml |
| Nuclease Free Water | 10 ml | 30 ml |
Storage and Stability
Carrier RNA should be stored at 2–8°C upon arrival. However, short-term storage (up to 12 weeks) at room temperature (15–25°C) does not affect their performance. The remaining kit components can be stored at room temperature (15–25°C) and are stable for at least 18 months under theseconditions
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