text: "Sequencing by synthesis" involves taking a single strand of the DNA to be sequenced and then synthesizing its complementary strand enzymatically. The pyrosequencing method is based on detecting the activity of DNA polymerase (a DNA synthesizing enzyme) with another chemoluminescent enzyme.  Essentially, the method allows sequencing a single strand of DNA by synthesizing the complementary strand along it, one base pair at a time, and detecting which base was actually added at each step.  The template DNA is immobile, and solutions of A, C, G, and T nucleotides are sequentially added and removed from the reaction.  Light is produced only when the nucleotide solution complements the first unpaired base of the template.  The sequence of solutions which produce chemiluminescent signals allows the determination of the sequence of the template.
  For the solution-based version of pyrosequencing, the single-strand DNA (ssDNA) template is hybridized to a sequencing primer and incubated with the enzymes DNA polymerase, ATP sulfurylase, luciferase and apyrase, and with the substrates adenosine 5´ phosphosulfate (APS) and luciferin.
  # The addition of one of the four deoxynucleotide triphosphates (dNTPs) (dATPαS, which is not a substrate for a luciferase, is added instead of dATP to avoid noise) initiates the second step. DNA polymerase incorporates the correct, complementary dNTPs onto the template. This incorporation releases pyrophosphate (PPi).
  # ATP sulfurylase converts PPi to ATP in the presence of adenosine 5´ phosphosulfate. This ATP acts as a substrate for the luciferase-mediated conversion of luciferin to oxyluciferin that generates visible light in amounts that are proportional to the amount. The light produced in the luciferase-catalyzed reaction is detected by a camera and analyzed in a program.
  # Unincorporated nucleotides and ATP are degraded by the apyrase, and the reaction can restart with another nucleotide.
  The process can be represented by the following equations:
  * PPi + APS → ATP + Sulfate (catalyzed by ATP-sulfurylase);
  * ATP + luciferin + O2 → AMP + PPi + oxyluciferin +  + hv (catalyzed by luciferase);
  where:
  * PPi is pyrophosphate
  * APS is adenosine 5-phosphosulfate;
  * ATP is adenosine triphosphate;
  * O2 is oxygen molecule;
  * AMP is adenosine monophosphate;
  *  is carbon dioxide;
  * hv is light.