Hi, Iím having troubles with my standard curve. I'm using DDCt method for relative quantification. Iíve already performed qPCR with other genes and had an excellent efficiency. But now Iím working with weakly expressed genes. Iíve already used both cDNA and purified PCR products, in 10x and 5x dilutions, the amplification plots seem to be perfect, regular distances between the dilution points, good correlation, but when I analyze the standard curve slope is between -4.20 and -4.08, and of course, low efficiency. There are no primers dimmers, no contamination cause NTC is OK. I donít know what is happening. Could somebody give me a suggestion? Thank you.