some gene families have relatively unique sequences for each gene, however there are many cases where pseudogenes have identical or almost identical sequences. the relatively short reads of NGS data complicates the situation in some areas of the genome. however when dealing with Eukaryotic RNA samples, you probably have to worry more about splicing variations and transcription initiation site variations..

the classical approach is to do mapping then go compare the results with the reference sequence.. but if you have a large coverage, i would go reading the alignment to check for differences..