1
Download HID EVOlution™ - qPCR/STR Setup System Getting Started Guide
Transcript
A Appendix A Troubleshooting Table 12 Troubleshooting qPCR/STR PCR results (continued) Observed problem Possible reason Suggested solution The sample IPC CT is higher than the IPC CT of the no template quantitation control (NTC) or of the quantitation standards (for example, if the sample IPC CT is approximately two CT greater than the NTC IPC CT or the CT of the standards). or Weak amplification (high CT value and low ΔRn value) of the human and/or male-specific targets and no or weak amplification of the IPC. • The DNA concentration is above 25 ng/µL. • Potential presence of PCR inhibitors. • If the DNA concentration is over 25 ng/µL, dilute the DNA extract, then requantify the sample. • If the DNA concentrate is below 25 ng/µL, or if the diluted DNA eluate still produces high IPC CT compared to the NTC or quantitation standards, repurify the sample. CE signal is too high Sample concentration is greater than the reported quantification result. 72 IMPORTANT! Repurification may result in the loss of additional DNA. Consider proceeding to amplification with a kit such as the AmpFlSTR® MiniFiler™ PCR Amplification kit, which is designed to obtain STR profiles from compromised samples such as those which may be inhibited and/or degraded. • Before the next run: – To ensure correct pipetting, clean and finger tighten the DiTi cones, diluter valves, and syringe fittings. Refer to the Tecan HID EVOlution™ - qPCR/STR Setup System Application Manual. – Confirm that you have sufficient system liquid, then flush the system and check for air bubbles and leaks. Refer to the Tecan HID EVOlution™ - qPCR/STR Setup System Application Manual. • Requantify (see IMPORTANT above). • Before the next run: – To ensure correct pipetting, clean and finger tighten the DiTi cones, diluter valves, and syringe fittings. Refer to the Tecan HID EVOlution™ - qPCR/STR Setup System Application Manual. – Confirm that you have sufficient system liquid, then flush the system and check for air bubbles and leaks. Refer to the Tecan HID EVOlution™ - qPCR/STR Setup System Application Manual. HID EVOlution™ - qPCR/STR Setup System Getting Started Guide
