Why is there always a problem with my reverse transcription?

1． You can test the integrity of RNA molecules. RNA quality has an important effect on cDNA synthesis results and it can be detected by agarose gel electrophoresis. Usually intact eukaryotic RNA should include two bright, well-margined bands, 28S and 18S, and the strength of larger bands should be about twice that of smaller bands, if the intensity is roughly the same.  

2. The OD value can be detected by the ultraviolet spectrophotometer to determine whether there is a product in the oligodt flip, so as to rule out whether there is a problem with reverse transcription. 

3． Judging from your description, CDNA is produced in reverse transcription, and the poor results of fluorescence quantification may not be a problem with the template, but there are problems such as reagent failure and experimental operation in the reaction system.