I got a slope of -3.9473 and r^2 value of 9.4 am worrying about its efficiency because I heard -3.32 is ideal slope .
Dear Raghavendra Basavaraja
Ideal R2 value is higher than 0.98 (0.99), and ideal slop is -3.314 (Efficiency= 1).
Please dilute your cDNA 10 million times using 10 fold series and perform RT-PCR.
you may not get the exact and ideal values but even then it does not mean that your primers are not fine. If you lack sufficient amount of cDNA....u can even got for primer dilution and draw the curve. Journals do accept little flexibility.
Please fins the attachment for details.
Alok
The slope depends on what dilutions of DNA you use. For example, if you use 1-to-10 serial dilutions and you expect your DNA to double each cycle, how many cycles does it take to make 10 times more DNA? The formula is 1*2^x=10, where x is the number of cycles. Hence x=log base 2 of 10. If you put this into Excel the formula looks like: "=LOG(10,2)". This gives you 3.321928...Conversely, if the you have 1-to-2 serial dilutions, the formula is 1*2^x=2, and x=1. This is the simplest case: you expect the slope of 1; it takes one cycle to double the amount of DNA.
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