Sequencing by synthesis refers to the process of sequencing one DNA strand and synthesizing, or combining, it with a complementary strand. The pyrosequencing protocol identifies the activity of a DNA synthesizing enzyme, called DNA polymerase, with another light emitting, or chemiluminescent, enzyme.
To sequence a single DNA strand, the pyrosequencing protocol synthesizes the complementary strand, base pair by base pair. Nucleotides are added and removed sequentially from the DNA template. Light generates only in cases where the first unpaired base of the DNA template is complemented by a particular nucleotide solution. Solutions that produce chemiluminescent signals enable the template sequence to be identified.
The pyrosequencing protocol is used in genome sequencing, or the determination or reproduction of an organism's genome sequence via scientific methods. Other applications for the pyrosequencing protocol include metagenomics, or the study of genetic material from environmental samples.