Having difficulties with your experiment?

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PCR is not perfect. Each step of the cycle should be optimized for each template and primer pair combination. The most important considerations for any PCR reaction are:

  • PCR enzymes—enzyme selection can affect the yield and specificity of PCR products. Get the basics on how to select the right PCR enzyme for your application.
  • Primer design—optimize your primer design using our custom oligo design tool.
  • Magnesium concentration—the correct magnesium concentration needs to be determined empirically for each primer and template combination you have; view this webinar on PCR optimization to help you determine the magnesium concentration.
  • Annealing temperature—the appropriate annealing temperature is based on the calculated Tm of your primers; watch a video on better than gradient to optimize your annealing temperature.

This can happen if the wrong plastics are used, or if the tubes are not being used with the correct tray/retainer set. Check the manual for directions on appropriate loading of the sample. Also, you can check for compatible plastics from here.

SimpliAmp Thermal Cycler

The heated lid is designed to make sure that optimized pressure is applied on the microplate for an efficient PCR reaction. When using PCR tubes, excess pressure on the tubes, from the heated lid can cause deformation of the tubes. To avoid this, we recommend using the Tray/Retainer set, as shown below:

  • Cat. No. 4381850, MicroAmp 96-Well Tray/Retainer Set – This Tray is for use with Single Tubes and/or Strip Tubes.
  • Cat. No. 4379983, MicroAmp 96-Well Tray – This Tray is for use with Single Tube with Cap and/or Strip Tubes with Caps.

ProFlex PCR System

First, make sure you are running the most up to date firmware. Newer firmware versions have fixed many wireless and mobile app issues. Firmware updates can be found here.

To set up the wireless connection in the ProFlex™ PCR System, please follow the steps outlined below:

  1. Power on the ProFlex™ Instrument, and in the Home screen, touch Settings.
  2. In the Settings screen, touch Device Settings.
  3. In the Device Settings screen, touch Network Connection. In the Network Connection Type screen, touch Wireless.
    The Search Wireless Network screen will come up and the wireless symbol will appear active. Touch Cancel to exit the screen.
    Note: During initial setup, if you selected the Wired option in the Network Connection Type screen, you will be required to enter your IP address if you selected the Static IP wired option. If you selected the Dynamic IP wired option, the IP address is automatically populated.
  4. Once a wireless connection has been detected, a list of the available networks will be displayed in the Choose Network screen. Touch the network name of your choice or touch Join Other Network.
  5. In the Find and Join a Network screen, enter the name and security type of the network.
    Note: When you touch in the Network Name field, a keypad will come up to facilitate making the entry.
  6. From the Security type drop-down menu, touch to select the security type and enter the relevant information in the screen.
    Note: Contact your IT Systems Administrator for information on security type.
    You can select from the following options:
    • WEP
    • WPA Personal
    • WPA2 Personal
    • WPA Enterprise
    • WPA2 Enterprise
  7. Touch Join to continue or Cancel to exit from the Find and Join a Network screen.
  8. Depending on the security type you have selected, enter the appropriate passwords and touch Join.
    If you entered the correct information, the Network Connection Complete screen will appear. Touch OK to continue. If you entered incorrect information, the Network Connection Failed screen will come up and you will be required to go back to security type screen to resolve the issue. Touch OK to continue.

Please follow these directions on how to set up mobile connectivity in the ProFlex™ PCR System with a mobile device.

  1. Connect your ProFlex™ PCR System to the internet using a wireless or wired connection.
    Note: See “How to set up the wireless connection in the ProFlex™ System” in Chapter 2 for setting up a wireless connection or refer to the ProFlex™ PCR System User Guide for setting up a wired connection in the ProFlex™ PCR System .
  2. Record the Instrument Serial Number and UUID number.
    • In the Home screen, touch Settings to open the Settings screen.
    • In the Settings screen, touch About Instrument.
    • In the About Instrument screen, touch About Instrument.
    • Record the Instrument Serial number and UUID.
  3. Turn on the connectivity to the Life Technologies™ server.
    • In the Home screen, touch Settings to open the Settings screen.
    • In the Settings screen, touch Device Settings.
    • In the Device Settings screen, touch Remote Service.
    • Using the On-Off toggle button, enable the Remote Service. Touch OK.

To set mobile connectivity in a mobile device, please follow the steps below:

  1. Download the "PCR Essentials Mobile Application" on your mobile device.
  2. Touch the "My ProFlex" button on the application.
  3. Enter your Lifetechnologies.com username and password.
    Note: If you do not have a Lifetechnologies.com username and password, register using the link on the mobile application.
  4. Enter the Serial Number of the ProFlex™ Instrument and add an instrument nickname if you want to.
    Touch Add Device.
  5. Once you have saved the instrument in your profile, the next time you log in, your instrument(s) will appear in a screen called "MY PROFLEXES"
  6. You can also add more instruments to your list by touching the +Add device button and repeat step 4 and step 5.

Make sure you have the most up-to-date firmware version, as this has fixed many screen-freezing issues. The latest firmware can be downloaded from here. If you cannot get access to the main menu, try turning off the machine, letting it sit for 1 minute, and restarting it without any block attached.

Veriti® Thermal Cycler

If the test fails:

  1. Verify that the:
    • Calibration of the TVK is up-to-date.
    • Probe tips are not cracked or damaged.
  2. Dispense 2 to 3 drops of mineral oil onto a cotton swab. Use the swab to apply a thin film of oil to the cone surface of each probe to improve heat transfer.
  3. Repeat the steps in “Before You Run 96-Well Instrument Tests” from the Multiprobe Temperature Verification Kit User Guide.
  4. Verify that the:
    • Probes are seated securely in the appropriate test wells.
    • Cables are connected securely to the ports.
    • Ribbon cable lies flat under the thermal cutout and is not crimped.
    • Heated cover is closed and locked down.
  5. Repeat the steps in “Run the Test” from the Multiprobe Temperature Verification Kit User Guide.

Please note, if the test fails again, contact Life Technologies at instrumentservices@lifetech.com.

GeneAmp® PCR System 9700

A guide to replacing lost calibration files for the GeneAmp® PCR System 9700 can be found here.

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