It depends substantially on what you mean by "DNA": for genomic DNA, where you can be reasonably confident that it

1) is genomic DNA, free of contaminating RNA or other nucleotides

2) it is the major source of DNA (not "genomic DNA plus other DNA from somewhere else" or whatever)

then yes, in theory you could normalise to DNA concentration. It's usually not difficult to find a suitable reference for genomic DNA, however, since most genes will be present in strict stoichiometry with number of genomes.

For cDNA, your DNA concentration isn't a useful measurement, as cDNA will contain many other nucleotide sources that will skew the reading (primers used for reverse transcription, leftover RNA, unincorporated nucleotides etc). Here the use of two or more reference genes in essential.

well

your amplification will depend on the state of your target, quality and quantity are not associated.

yes, the reference gene is better used. the delta delta Ct method is the best way to run RT-qPCR protocols, just take some time and follow this link (https://toptipbio.com/delta-delta-ct-pcr/) to prevent false results and ensure reliable data.

all the best

fred

In theory, yes. In practice, yes (with a LOT of caveats).

If you are using DNA as your template (NOT cDNA)

If you are measuring gene copy number (NOT gene expression)

and

you have a way to know EXACTLY how many cells you started with

and

your DNA extraction protocol is very consistent AND very efficient

then it's possible.

One example I know of is in C. elegans nematodes. The larvae have a known number of cells. You extract DNA from EXACTLY one (or two, or some other number) of worms. Then you can do things like measure mitochondrial genome copies per genome.

Good luck!