I am planning to purchase a cell disruptor/ homogenizer, probably the kind that shakes beads (steel, glass, or ceramic) at very high speeds, to break open plant leaf (and possibly wood or seeds) cells. I will then be extracting DNA for sequencing, genotyping, etc. I hope to obtain high molecular weight DNA for microsatellite, RADseq, SNP genotyping etc. There are so many options out there for this piece of equipment. Some colleagues use a mortar and pestle, but that is really not enough throughput for me- I'd like to do at least 12 samples at a time. Could you please share your preferred equipment, what quality/ quantity/ mol weight of DNA you get from it, how fast and easy to use it is, if it has a cryo option, and any other suggestions (including tips on extractions)!
The options I have looked at are:
Fastprep (http://www.mpbio.com/fastprep5G.php?country=223)
Beadbeater (http://www.biospec.com/product/34/mini_beadbeater/)
miniG (http://www.spexsampleprep.com/minig)
Retsch mill (http://www.retsch.com/en/products/miling/ball-mills/mixer-mill-mm-400/function-features)
Disruptor Genie (https://us.vwr.com/store/catalog/product.jsp?catalog_number=89202-318)
Bead Ruptor (https://us.vwr.com/store/catalog/product.jsp?catalog_number=10032-378)
VWR Homogenizer (https://us.vwr.com/store/catalog/product.jsp?catalog_number=12621-164)
Precylls (http://homogenizers.net/collections/bead-mill-homogenizers/products/precellys-evolution#review)
Hi Sean,
We are using a Retsch mill with SS balls. its fast and yeilds good quality DNA usually, depending upon the quality of the leaves. we usually run the samples at 25 Hz for 2-3 mins in the prec-cooled sample tube holder.
Best wishes,
SD
Thank you very much SD. Do you cool the tubes in liquid nitrogen? Also what type of DNA genotyping do you do with your samples (RAD-seq, microsats, AFLP, genome sequencing)? Thanks!
Dear Sean,
we generally precool the tube adapter by keeping them in -20C for 30 mins. We do PCR for viral genome amplification and sequencing. By using the DNA obtained by this method we successfully amplify upto 1.4 kb of ampilcons.
https://www.qiagen.com/pk/shop/automated-solutions/sample-disruption/tissuelyser-ii#orderinginformation
I've always used the BioSpec Beadbeater (http://www.biospec.com/product/34/mini_beadbeater/). They are cheap and work really well. Only thing is that for hard plant tissue (needles and wood) you must use steel beads and reinforced micro vials (http://www.biospec.com/product/462/xxtuffreinforced_microvials/).
add plant and beads to vials, dive them on liquid nitrogen for 30 sec and shake on high frequency for 30 sec, do it two or three times depending of the hardness of the tissue and you get a fine powder ready for dna extraction.
Hi Sean!
I've been using a Retsch Mill for high throughput (2x96 plates) with great success for everything from single gene sequencing to NGS. I've used the fancy Qiagen tungsten carbide beads and cheap stainless steel from Amazon (ball bearings) without any detectable difference. With algal material, we haven't been freezing in liquid nitrogen, just grinding dried material.
We found a good mill on eBay that is much cheaper than the Retsch or Qiagen new models.
Let me know if you have anything other questions!
Stacy
The plant people in our center use the Fastprep and the Lysis Matrix D from MP Bio for plant tissue but they use the metal beads for wood pieces.
I've also heard that you can clamp your samples down to a paint shaker and use that.
Thanks Sheina, Stacy and Rafael! I was thinking of the Beadbeater or the Retsch. The Beadbeater is a third of the price of the Restch which sounds nice (!), and I couldn't tell if there was any difference between the machines. Rafael, do you freeze dry or silica dry your material, or just use fresh? Thanks all!
I used silica because I needed to ship my samples overseas, and it would have been much harder to get permits to ship samples in liquid preservation buffers. Fresh only if you can run the extraction right after collecting the sample.
Silica + freezer allow you to keep your samples viable for several years. A -80 C freezer can probably keep your samples good for gen work for the duration of your scientific career!
Sean,
I don't know if you can use eBay, but here is one on eBay. Have a look around as there's others.
We contacted the seller and it turned out to be a company. They let us buy it through their company website. We've had no problems and it's gotten loads of use.
stacy
http://www.ebay.com/itm/Retsch-TissueLyser-Mixer-Mill-Grinder-Homogenizer-Cell-Disrupter-Retsch-MM200-/252228227514
Thanks everyone for the answers so far! Do share this with your colleagues to see if we can get some more insight!
Hi Sean,
You might want to consider the Bullet Blender Gold as well. Great throughput, very simple dry ice cooling method, and a very good price.
https://homogenizers.net/collections/bead-mill-homogenizers/products/bullet-blender-gold
A Disruptor Genie would have no chance of getting through wood or seeds. It's just not powerful enough.
Also, that "VWR Homogenizer" that you listed is just a Talboys High Throughput Homogenizer that they slapped their name on and marked up. https://homogenizers.net/collections/bead-mill-homogenizers/products/talboys-high-throughput-homogenizer
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