Monoubiquitylation of Histone H2B in Giardia Lamblia

Faculty Mentor(s)

Dr. Michael Pikaart, Hope College
Dr. Aaron Best, Hope College
Dr. Dereje Desta, Hope College

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Covalent modifications of core histone proteins are essential to gene transcription. One such modification is the monoubiquitylation of H2B, a process which has been shown to occur in all tested eukaryotes. However, this process has not been studied in Giardia lamblia, which is thought to be the earliest-diverging eukaryote. To create gene constructs to test for the monoubiquitylation of histone H2B, histone H2B was amplified from Giardial genomic DNA. Antibody tags were attached to the gene through PCR primer design. After cloning the gene it was ligated into pNMyb, a plasmid containing a Giardial promoter and origin of replication. Subsequently the plasmid was introduced into G. lambalia through electroporation. Western blotting, RT-PCR, and Immunoflourescence assays have indicated low levels of HA-tagged H2B. The expression vector is being remade, and G. lambila will be transfected with the new expression vector.

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