You are performing a biochemical purification of enzymes involved in DNA replication. You have purified the replisome. You wish to purify the primosome. You perform further separation techniques on your purified replisome. How will you test to determine that you have purified the primosome

Confirm that the sample has both helicase and primase activities, but not the ability to synthesize DNA

            1. Helicase enzymes that unwind and separate the DNA strand

            2. Replication factor C (RFC), a DNA-dependent ATP that acts as an activator of DNA polymerase.

           3. Proliferating cell nuclear antigen (PCNA)

           4. DNA gyrase/topoisomerase that cuts and reseals DNA strands, which is essential for DNA synthesis.

           5. Primase enzyme that synthesizes short RNA fragments called primers.

          6. DNA polymerase III, an enzyme that catalyzes the polymerization of deoxyribonucleotides into a DNA strand.

           7. DNA ligases enzymes that form phosphodiester bonds at a single-strand break in DNA

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step 1: nitrogen fixation- conversion of atmospheric no₂ into forms useful to plants by lightning, bacteria, and cyanobacteria.

step 2: nitrification-process of converting ammonia to nitrite or nitrate.

step 3: assimilation-plant roots absorb new form of nitrogen.

step 4: ammonification-conversion of organic nitrogen to ammonia by bacteria involved in decomposition.

step 5: denitrification-bacteria turns the decomposed matter back into n₂o, which is released into atmosphere; repeat cycle.