Chromatin is the essential part of transcription mechanism inside the cell. However, when native chromatin templates were transcribed in vitro with yeast whole cell extract, very poor or no transcription was observed. This indicated that an integral part(s) of transcription machinery essential for transcribing chromatin template either was missing or inactivated in the whole cell extract. My research project in Dr. Roger Kornberg's lab is to develop a novel system to study the mechanism of transcription on chromatin templates. This novel transcription system is also called """"""""permabilized cell"""""""" system. In this system, yeast spheroplasts are permabilized by fast freezing and thawing (1). This gentle treatment of the spheroplasts which allows the large protein molecules, e.g., transcription factors, to diffuse into the cell, causes the least disruption of the cell functional systems. For example, after the treatment, all the organelles remain intact and are capable of protein transport and processing (1). This system is excellent for studying transcription of chromatin templates. I am currently trying to understand the induction of genomic Gall gene using the cells grown under repressed conditions by adding externally, e.g., the known transcription activator, Ga14-VP16. I am asking the question what factors are needed to reconstitute the level of transcription of Gall gene seen under galactose induced conditions.
