Abstract: Disclosed are methods for determining one or more nucleotides at one or more nucleotide positions of a polynucleotide sample, the polynucleotide sample comprising heterogeneous polynucleotides having different nucleotides at the nucleotide positions. The disclosed, methods may be utilized to control for sequencing bias during sequencing of the polynucleotide sample. Suitable samples may include patient samples for use in diagnosing, prognosing, and treating the patient.