eLabNotebook > Nucleic Acid Prep & Purification > RNA Purification > Promega MagneSil*
Total RNA mini-Isolation Biomek® FX


Promega's MagneSil* Total RNA mini-Isolation System on the Biomek® FX

Promega Corporation – www.promega.com

For additional information try our Maintenance and Troubleshooting section or post a question to our Biomek User Forum.  New members can subscribe to our User Forum here.

Click here to download a printable PDF version of the following:

Problem Possible Cause Solution
Low RNA yield
  • Initial sample mass too large.
  • The MagneSil* Total RNA mini-Isolation System is optimized for purification from =1 × 105 cells, =2mg tissue lysate in 100µl, or =20µl whole blood. Exceeding these limits will significantly reduce both yield and concentration of the purifed RNA and will cause excessive clumping of MagneSil* RNA Paramagnetic Particles, making handling of the particles difficult.
Low RNA yield
  • Use of sample lysate that has been stored at ?20°C or ?70°C. Lysate samples that have been frozen may have a decreased amount of total RNA.
  • For optimal performance, purify the total RNA as soon as the lysate is prepared.
Low RNA yield
  • Sample lysates have undergone multiple freezethaw cycles. Samples that have been frozen and thawed repeatedly will have some RNA degradation.
  • Use fresh samples whenever possible.
Low RNA yield
  • Tissue culture cells low in total RNA. The yield of total RNA may vary depending on the sample type.
  • If total RNA yields are low, increase the amount of starting material processed.
Low RNA yield
  • RNA Lysis Buffer not added to tissue lysates.
  • Make sure that RNA Lysis Buffer is added to all sample lysates.
Low RNA yield
  • Steps not followed correctly, or the wrong reagents were used.
  • Use the MagneSil* Total RNA mini-Isolation System reagents in the order specified. This ensures that the RNA remains bound to the MagneSil* RNA PMPs during the purification process.
Low RNA yield
  • Ethanol not added to DNase Stop Solution.
  • Ensure that ethanol is added to the DNase Stop Solution. See Section IV.A.
Low RNA yield
  • Incorrect concentration of ethanol wash. Ethanol concentrations lower than 90% will result in reduced yields.
  • Make 90% ethanol wash solution fresh before each purification.
Low RNA yield
  • Failure to resuspend MagneSil* RNA PMPs in reagent bottle.
  • Thoroughly resuspend MagneSil* RNA PMPs in the reagent bottle before dispensing to the sample processing plate to ensure even distribution of particles.
Low RNA yield
  • Inaccurate dispensing of MagneSil* RNA PMPs. The volumes of MagneSil* RNA PMPs used in the protocol are optimized for yield and purity of RNA.g
  • Use of 25% greater than or less than the recommended amount will result in decreased yield (e.g., purification from 1 × 105 cultured cells requires 30µl of MagneSil* RNA PMPs; no less than 22.5µl and no more than 37.5µl of particles should be used per isolation.)
DNA contamination
  • DNase Solution not prepared correctly.
  • For each plate purification 5.2ml Yellow Core Buffer, 575µl MnCl2 and 275µl of DNase I are mixed to make a DNase Solution.
DNA contamination
  • Insufficient incubation with DNase Solution.
  • Incubate DNase Solution for at least 10 minutes. DNase incubation time can be lengthened up to 20 minutes.
DNA contamination
  • DNase Solution stored or frozen before use.
  • Make DNase Solution fresh before each use. It cannot be prepared and then stored.
MagneSil* RNA PMPs clump
  • Too much sample material used.
  • MagneSil* RNA mini-Isolation System input limitations are =1 × 105 cells, =2mg tissue lysate in 100µl volume and =20µl whole blood
MagneSil* RNA PMPs clump
  • Lysate too concentrated.
  • If the lysate is too viscous, dilute with RNA Lysis Buffer until it becomes easier to pipette.
MagneSil* RNA PMPs clump
  • Insufficient mixing.
  • Vigorously mix during wash and incubation steps to resuspend the MagneSil* RNA PMPs.
RNA degradation
  • RNase introduced during purification/handling.
  • Use RNase-free plastic- or glassware during the purification process. Use filter tips during all pipetting steps. Wear gloves at all times. RNases introduced after elution will degrade RNA.

* All trademarks are the property of their respective owners. Where applicable, the PCR process is covered by patents owned by Roche Molecular Systems, Inc., and F. Hoffman-LaRoche, Ltd.

eLabNotebook Sitemap

©1998 - 2006 Beckman Coulter, Inc.