Hi everyone, I have performed RT-PCR with cDNA recently, but may I know why I got this kind of amplification plot even for the housekeeping gene??

Maw Shin (Gareth) Sim @Maw-Shin-gareth-Sim

14 November 2022 1 10K Report

For your information, I am using ABI StepOne Plus machine. The housekeeping gene is Beta-actin.

However, the results showed its melting curve and fluorescence plot look good.

Appreciate it if any experts here could help.

Your amplification plot indicates that your PCR reaction did not work at all. This called suboptimal qPCR amplification curves. You need to optimize the amount of cDNA you are using as a template and check your Mastermix. Also, make sure you are using the correct PCR parameters that are compatible with your primers melting points.

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