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Biomiga Bacterial RNA Kit - R6616-01

  • Model: R6616-01
  • Manufactured by: Biomiga

£128.00

Description :

Introduction

The EZgeneTM total RNA kit provides an easy and fast method for isolating total RNA from Gram-positive (B. subtilis) Or Gram-negative (E. coli) Bacteria 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.

Storage and Stability

DNase I (optional) and lysozyme should be stored at -20℃. All other components can be stored at room temperature. All kit components are guaranteed for 12 monthsr from the date of purchasing.

Kit Contents

Catalog#

R6616-00

R6616-01

R6616-02

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 dH2O

500 µL

10 mL

30 mL

DNase Stop Buffer

200 µL

2.4 mL

12 mL

ezBind Columns

4

50

250

Collection Tubes

8

100

500

Lysozyme

1.2 mg

15 mg

75 mg

User Menu

1

1

1

Note:DNase I are not supplied. They could be purchased from Biomiga.

Before Starting

Prepare all components and get all necessary materials ready by examining this instruction booklet and become familiar with each steps.

Important

  •  Add 1 % volume of β-mercaptoethanol to Buffer LY before use and store at 4 ℃.
  •  Add 8 mL(R6616-00) or 80 mL (R6616-01) or 96 mL (R6616-02) 100% ethanol to RNA Wash Buffer before use.
  •  Add 800 µL(R6616-00) or 9.6 mL (R6616-01) or 48 mL (R6616-02) 100% ethanol to DNase Stop Buffer before use.
  •  Prepare a lysozyme stock solution at 3 mg/mL or 0.4 mg/mL with Elution Buffer or TE Buffer and aliquot into adequate portions. Store each aliquot at -20 ℃ and thaw before use.

Materials supplied by users

  •  Tabletop microcentrifuge and 1.5 mL sterile tubes.
  •  Vacuum manifold if use vacuum protocol.
  •  100% ethanol
  •  Optional: DNase I, DNase Buffer

Note: Perform all steps including centrifugation at room temperature

Trouble Shooting Guide

Problem

Possible reason

Suggested Improvement

Low A260/A280 ratios

Protein contamination

Do a Phenol:Chloroform extraction. Loss of total RNA (up to 40%) should be expected.

Guanidine Thiocyanate contamination

Add 2.5 volumes of ethanol and 0.1M NaCl (final concentration) to precipitate RNA. Incubate for 30 min at -20 ℃. Centrifuge at 10,000 g for 15 min at 4 ℃. Resuspend the RNA pellet in DEPC-treated water.

Low Yield

RNA in sample degraded

Freeze samples immediately in liquid nitrogen and store at -70 ℃ after collect it.

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.

Ethanol not added to buffer

Add ethanol to the RNA Wash Buffer and DNase Stop Buffer 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.

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 1-2x106 or increase buffer volume and do multiple loadings to column.

Operating Protocol :

r661633521_448


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