StepOnePlus is a Step Above the Rest

April 11, 2014

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Penn State University
Biobehavioral Health
Lab Manager

Company:

Life Technologies

Product Name:

StepOnePlus

Catalog Number:

4376600

Research in our lab focuses on how temperament or personality relates to physiology and health. The larger goal is to determine how and why individuals differ in their behavioral and physiological responses to common challenges and to understand how this influences individual susceptibility and/or resilience to certain disease states and rates of aging. We use animal models for much of our work and look at gene expression levels, mainly in the brain, that are up-regulated or down-regulated in relation to different behavioral types. We chose the StepOnePlus due to its association with Life Technologies high quality (albeit expensive) products, its ease of use, helpful technical staff, and interactive, colorful software program.

That said, there are a few goofy issues with the machine that have been discovered over time with lots of use. 1) if the main screen shows a triangle with an exclamation point right after starting a run, stop immediately. This happens rarely and we have not been able to figure out why but, if you continue to run the plate, you will not be able to collect the results at the end of the run. Opening the template and renaming the file slightly usually resolves this problem and you can proceed with the run. 2) The machine allows you to program a 4 degree hold but it cannot carry this out and if put into your program, you will not be able to collect results at the end of the run.

Experimental Design and Results Summary

Application

Comparative qPCR

Starting Material

Rodent brain tissue

Protocol Overview

Tissue is homogenized with TRI reagent and then RNA is extracted and reverse transcribed to cDNA with a high capacity reverse transcriptase kit and then taqman assay components were used to run the standard qPCR fo several different genes of interest using beta actin as the housekeeping gene for comparison.

Tips

The kit says to prepare on ice in the dark but both of these are unnecessary. The probe should not be exposed to a lot of light for a long period of time but the amount of time it takes to set up a plate is not enough to affect the product. Make sure to aliquot the probe when it first arrives to avoid freeze thaws. Also, the master mix has a short shelf life of about 6 months and is very expensive so make sure not to order more than you will use in that time.

Results Summary

Our results are proprietary but we use the delta delta CT method for our analyses. Basically, this is derived by getting the delta CT (the amount of our gene of interest minus the amount of housekeeping gene in a sample) and then the delta delta CT (the delta CT of your experimental sample minus the delta CT of your control sample) and then your relative gene expression equals 2^-delta delta CT.

Features Summary

The machine does all of these calculations for you but this only works if your entire experiment can be run on one plate. If you have to use multiple plates, like me, I only get the delta CT values from the software and calculate the rest by hand.

Additional Notes

None

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Summary

The Good

Easy to use, great for visual people, good results

The Bad

Goofy software issues; expensive

The Bottom Line

If looking for a machine to do quality PCR analysis and can afford to purchase Life Technology supplies, this machine is a nice option.

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