Zymo Plasmid Midiprep Kit - Good Performance At A Reasonable Price

October 19, 2016

Duke University
Immunology
Graduate student

Overall

Quality of Results

Ease-of-Optimization

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Company:

Zymo Research

Product Name:

ZymoPURE™ Plasmid Midiprep Kit

Catalog Number:

D4200

I have always used Qiagen's HiSpeed Midi kits to perform plasmid midipreps, but wanted to try another supplier's kit. I requested a sample Midiprep kit from Zymo, which was free and shipped quickly. The Zymo protocol has fewer steps than Qiagen's, and seems to take ~50% less time than the Qiagen protocol. Overall, the Zymo kit components are well-designed, although one piece is too big to fit in standard size conical tube holders.The prepped plasmid DNA is highly pure (as measured by Abs spectroscopy and agarose gel electrophoresis), and should be suitable for transfection of retrovirus packaging cell lines.

Experimental Design and Results Summary

Application

Plasmid DNA midiprep

Starting Material

E. coli cell pellet from 50 mL overnight culture

Protocol Overview

The kit follows a standard cell lysis and neutralization protocol using NaOH/SDS and KOAc. The first three buffers are color-coded -- this (1) prevents the user from confusing the bottles, and (2) makes it very easy to determine when the solutions have been fully mixed after each step. The neutralized lysate is clarified by forcing the liquid through a filtered syringe. The clarified lysate is then passed through a DNA-binding column matrix, either by vacuum manifold or by centrifuge (swing-bucket type capable of holding 50 mL conical tubes). The column is then washed 3 times in a tabletop microcentrifuge (or vacuum manifold), and the DNA eluted in 100-200 uL of H2O/buffer.

Tips

After lysate neutralization, the crude lysate is poured into the filter syringe and incubated (upright) for 5 min. The syringe is too large to fit in a standard size 50 mL conical tube rack, so one must either use another (bigger) rack, hold the syringes upright in one's hands, balance the syringes on the benchtop (it works, but it's dangerous), or just skip the incubation period altogether (it actually doesn't seem necessary).

Results Summary

The eluted plasmid DNA was highly pure (high A260/A280 and A260/A230 ratios). Gel electrophoresis confirmed pure, supercoiled plasmid and negligible genomic DNA contamination. The yield was lower than I anticipated (I got 100 ug per plasmid, I expected ~250 ug/plasmid based on preliminary minipreps, Zymo says 300 ug is the max yield for this kit), but comparable or slightly better than what I was getting with Qiagen's kit.The protocol took me 1 hour for two plasmids. It would probably take 90-120 min with Qiagen's kit, and I would be sweaty by the end, too.

Additional Notes

None

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Summary

The Good

Great final purity, easy to use, love the color-coded buffers, and Zymo's products typically have lower-than-average prices

The Bad

Lower-than-expected yield, filter syringe not properly sized to fit in standard laboratory tube racks.

The Bottom Line

I like the convenience and performance:price ratio. The improperly sized filter syringe isn't a deal-breaker, and the underwhelming yield might be improved with optimization of E coli culture conditions. The ability to use standard centrifuges for the column loading/wash/elution steps is a big improvement over the gravity-based or syringe-based steps used in other supplier's kits (I'm looking at you, Qiagen).I like the kit enough to try to persuade our lab manager to start stocking the Zymo Midi kit.

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