Product Description
Buffer LY, RB, RNA Wash Buffer, DEPC-Treated ddH2O, ezBind Columns, Collection Tubes, DNase I (Optional), User Menu
Features :
Tissue,Eukaryotic cells RNA purification miniprep Kit: Purify up to 100μg total RNA from 5×106 Eukaryotic cells or 30mg tissue
Introduction
The EZgeneTM Tissue RNA Kit provides an easy and fast method for isolating total RNA from tissues, cultured cells within 30 min. Only trace genomic DNA exists in the purified RNA, which can be eliminated by DNase I treatment (See detail in the protocol) when it is necessary. This kit purifies up to 100 µg of total RNA from eukaryotic cells or animal tissues. The purified RNA is ready for RT-PCR, Northern blotting, polyA+ RNA purification, nuclease protection, and in vitro translation.
Storage and stability
DNase I (opsional) harus disimpan pada -20 ° C. Semua komponen lainnya dapat disimpan pada suhu kamar. Semua komponen kit dijamin selama 12 bulan sejak tanggal pembelian.
Kit contents
| Catalog# |
R6311-00 |
R6311-01 |
R6311-02 |
| Preps |
4 |
50 |
250 |
| Buffer LY |
2.4 mL |
28 mL |
135 mL |
| Buffer RB |
3 mL |
30 mL |
135 mL |
| RNA Wash Buffer |
2 mL |
20 mL |
3 x 24 mL |
| DEPC-Treated ddH2O |
500 µL |
10 mL |
30 mL |
| ezBind Columns |
4 |
50 |
250 |
| Collection Tubes |
8 |
100 |
500 |
| DNase I (Optional)* |
25 u |
300 u |
1500 u |
| DNase Stop Buffer |
200 µL |
2.4 mL |
12 mL |
*Solutions are not supplied. They could be purchased from Biomiga.
Safety information
Buffer LY mengandung garam chaotropic, yang dapat membentuk senyawa reaktif jika digabungkan dengan pemutih. Jangan menambahkan pemutih atau larutan asam langsung ke limbah persiapan, kenakan sarung tangan dan kacamata pelindung saat menangani.
Before starting
Persiapkan semua komponen dan persiapkan semua bahan yang diperlukan dengan memeriksa buklet instruksi ini dan menjadi terbiasa dengan setiap langkah.
Important
Tentukan volume Buffer LY yang akan digunakan dan tambahkan 20 µL b-mercaptoethanol (b-ME) per 1 mL Buffer LY sebelum digunakan. Buffer LY berisi b-ME dapat disimpan pada suhu ruangan hingga 1 bulan.
Kristal dapat terbentuk di Buffer LY, larutkan endapan pada suhu 37 oC sebelum digunakan.
Tambahkan 8 mL (R6311-00) atau 80 mL (R6311-01) atau 96 mL (R6311-02) etanol 100% ke RNA Wash Buffer sebelum digunakan. Etanol akhir adalah 80% (v / v).
Opsional: Tambahkan etanol 100% 800 µL atau 9,6 mL (R6311-01) atau 48 mL (R6311-02) ke DNase Stop Buffer sebelum digunakan. Etanol akhir adalah 80% (v / v).
Materials supplied by users
- Tabletop microcentrifuge and 1.5 mL sterile tubes.
- Vacuum manifold if use vacuum protocol.
- 100% ethanol.
- b-mercaptoethanol.
- Optional: DNase I, DNase Buffer
Note: Perform all steps including centrifugation at room temperature
Trouble Shooting Guide
| Problems | Possible reasons | Suggested Improvements |
| Low A260/A280 ratios
|
Protein contamination | Do a Phenol:Chloroform extraction. Loss of total RNA (up to 40%) should be expected. |
| Low A260/A280 ratios
|
Guanidine Thiocyanate contamination
|
Add 2.5 volumes of ethanol and 0.1M NaCl (final concentration) to precipitate RNA. Incubate for 30 min at -20°C. Centrifuge at 10,000 g for 15 min at 4°C. Resuspend the RNA pellet in DEPC-treated water. |
| Low Yield | RNA in sample degraded
|
Freeze samples immediately in liquid nitrogen and store at -70°C after collect it. |
| Low Yield | The binding capacity of the membrane in the spin column was exceeded | Use of too much tissue sample exceeding the binding capacity of spin column will cause the decreasing of total RNA yield. |
| Low Yield | Ethanol not added to buffer | Add ethanol to the RNA Wash Buffer and DNase Stop Solution before purification. |
| Genomic DNA contamination
|
Too much total RNA sample was used in RT-PCR. | Reduce total RNA amount used in RT-PCR to 50-100 ng. |
| Genomic DNA contamination
|
The sample may contain too much genomic DNA.
|
Reduce the amount of starting tissue in the preparation of the homogenate. Most tissues will not show a genomic DNA contamination problem at 30 mg or less per prep.
Reduce cell numbers to 1-2×105 or increase buffer volume and do multiple loadings to column. |
MORE INFO :
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No HP : 0813 1754 7882
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