Observe the Log View of the amplification plots. Manually define the Threshold Value (Figure 3): i. *IMPORTANT: Ensure that all selections remain consistent across all PCR Array runs in the same analysis. Noise Slope Correction improves the data when raw data backgrounds are observed to slope upward or downward before the takeoff point (C t). Selection of this option can improve data whose baseline (initial cycles) is noticeably sloped. (Optional) Select Noise Slope Correction. Thus, better results may be achieved if the initial cycles are ignored. Fluorescent signal from the initial cycles may not be representative of the remainder of the run. Select Dynamic Tube (default analysis setting) to ensure the average background of each well is determined just before amplification commences. Observe amplification plots in Linear View.
Calculate the threshold cycle (C t) for each well using the instrument s software.
Under Quantitation tab, select Cycling A. Once the PCR run is complete, observe the Sample Bank (right side of screen, Figure 2). Confirm Profile tab, verify desired profile. Rotor Selection tab, select Rotor-Disc Ensure locking ring has been attached to the Rotor-Disc 100 and check Locking Ring Attached 9.
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Open the Rotor-Gene Q Series Software Under the New Run dialog box, click on the Quick Start tab, and select Open a Template In Another Folder. If the Rotor-Gene Q is off, switch on the instrument, and ensure the standby light is lit. of 5Ģ Figure 1 Edit Profile Tab Performing Real-Time PCR Detection 1. Click Save Template and enter RT2_Rotor_Gene_Q as the template name. Ensure Perform Optimisation Before 1st Acquistion is checked. Under the Auto-Gain Optimisation Setup tab, click Optimise Acquiring. Ensure Optimize gain before melt on all tubes is checked. Under the Edit Profile tab (Figure 1), adjust parameters to reflect the following: Hold Hold Temperature: 95 C Hold Time: 10 mins 0 secs Cycling This cycle repeats 40 time(s) 95 C, 15 seconds, Not Acquiring 60 C, 30 seconds, Acquiring to Cycling A on Green Click Insert After New Melt. Under the Miscellaneous Options tab, set Reaction Volume (µl) to Click Next. Under the Welcome to the Advanced Run Wizard! tab, select Rotor-Disc Ensure locking ring has been attached to the Rotor-Disc 100 and check Locking Ring Attached 7. The New Run dialog box will appear (Note: the New Run dialog box may open automatically). Open the Rotor-Gene Q Series Software 2.0 on the desktop of the computer that is connected to the Rotor-Gene Q.
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For more information, please click the help button or refer to the Rotor-Gene Q Manual available at Set up a PCR protocol template file on the Rotor-Gene Q Series software as follows: 1. 1 QIAGEN Rotor Gene Q: Software V2.0 Instrument Setup Instructions for RT 2 Profiler PCR Arrays Preparation Before the Experiment (Presetting the Machine will save time for your run): Please make note of the Rotor-Gene Q Series software version on your instrument.
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