morgan@go2biotech.comDP412-RNAclean Kit
RNAclean Kit
For RNA cleanup and concentration with small elution volumes
RNAclean Kit (Spin Column)
Cat. no. 4992728
Storage
RNAclean Kit should be stored dry at room temperature (15-30°C) and are
stable for 15 months under these conditions.
Introduction
The kit combines the selective binding properties of a silica-based
membrane with the speed of microspin technology. RNA could be adsorbed
to silica-based membrane efficiently and exclusively in the solution under
high salt concentration. At the same time, protein, saline ions, organic
impurities and other contaminants could be removed to a large extend.
RNA will be eluted in the solution under low salt concentration. Up to 20
μg RNA sample could be processed each.
The kit is used for RNA purification from enzymatic reaction (such as DNase
digestion, Protease treatment, RNA labeling and so on). The purified RNA
is free of protein contamination, and can be used in many downstream
experiments such as Northern blot, Dot blot, mRNA extraction, synthesis
of cDNA, primer extension, differential display, etc.
Notes for preventing RNA contamination
- Wear gloves when handling RNA and all reagents, as skin is a common
source of RNase. Change gloves frequently. - Use sterilized plastic ware and tips to avoid cross-contamination.
- Plastic or glass ware should be RNase-free. To wipe off RNase, the
glassware could be dried at 150°C for 4 hours, while plastics could be
dipped in 0.5 M NaOH for 10 min, and washed by RNA-Free ddH2O
thoroughly and sterilized. - Prepare the reagents with 0.01-0.1% DEPC Treated RNase-Free ddH2O.
Treat microcentrifuge tubes and tips with DEPC and autoclave to remove
any trace of DEPC before use.
Protocol
β-mercaptoethanol (β-ME) should be added to Buffer RK for the first use.
Add 10 μl β-ME per 1 ml Buffer RK. Please prepare the solution fresh
right before use. Buffer RK containing β-ME can be stored at 2-8°C for up
to 1 month. Redissolve it before use when precipitate is formed.
Ensure that 96-100% ethanol is added to Buffer RW as indicated on the
tag of bottle.
All the steps as below should be performed in a ice bath.
- Adjust the sample to a volume of 100 μl with RNase-free water. Add 350μl Buffer RK(ensure β-ME is added before use), and mix well.
- Add 250 μl 96-100% ethanol to the diluted RNA, and mix well by
pipetting. Do not centrifuge. Proceed immediately to step 3. - Transfer the sample and precipitate to a Spin Column CR2 placed in a 2 ml collection tube . Close the lid gently, and centrifuge for 30 sec at
12,000 rpm. Discard the flow-through. - Add 500 μl Buffer RW(ensure that ethanol is added into Buffer RW)to
the spin column. Close the lid gently, and centrifuge for 30 sec at 12,000
rpm to wash the spin column membrane. Discard the flow-through. - Repeat step 4.
- Centrifuge for 5 min at 12,000 rpm and discard the flow-through.
- Place the Spin Column CR2 in a new 1.5 ml collection tube. Add 14-20 μl
RNase-Free ddH2O directly to the center of the spin column membrane.
Incubate for 2 min at room temperature. Close the lid gently, and
centrifuge for 2 min at 12,000 rpm to elute the RNA.
Note: Elution buffer volume should be over 14 μl, otherwise RNA yield
and quality will be reduced.
The pH of the elution buffer has great effect on elution efficiency. The
pH should be at the range of 7.0-8.5 if water is used as elution buffer,
otherwise, efficiency of elution will be reduced. RNA product should be
stored at -20°C. - It is recommended to repeat step 7 for higher yield , then combine the
eluate .
DP412-RNAclean Kit
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This product is for scientific research use only. Do not use in medicine, clinical treatment, food or cosmetics.
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