Event Title
Search for the Nuclear Localization Signal in the Yeast mRNA Methyltransferase, IME4
Faculty Sponsor
Mary Clancy
College(s)
College of Sciences
Submission Type
Poster
Description
The IME4 gene in S. cerevisiae (yeast) codes for a nuclear proteins that is responsible for the methylation of mRNA in the nucleus. Equivalent proteins are found in most eukaryotic cells. However, the IME4 protein does not contain any previously identified nuclear localization segment (NLS) found in other genes; through this experiment we hope to be able to identify the portion of the protein that serves this function. Based on previous experiments in which segments of the IME4 gene were removed without interfering with nuclear localization, we hypothesize that the NLS is in the second half of the protein (C-terminal half). In order to test this idea, we attached the segment of IME4 that codes for the C-terminal half to a plasmid that encodes green fluorescent protein (GFP); because GFP does not undergo nuclear localization naturally, if GFP is found in the nucleus our hypothesis will be supported. To carry out this process, we first synthesized the modified copies of IME4 by amplification using the polymerase chain reaction (PCR). Then, we attached the IME4 gene to plasmids encoding GFP; these plasmids were then evaluated using agarose gel electrophoresis to determine whether they were formed correctly. Once the plasmids were confirmed, they were transformed into S. cerevisiae cells. Currently, the location of GFP is being examined using fluorescence microscopy.
Search for the Nuclear Localization Signal in the Yeast mRNA Methyltransferase, IME4
The IME4 gene in S. cerevisiae (yeast) codes for a nuclear proteins that is responsible for the methylation of mRNA in the nucleus. Equivalent proteins are found in most eukaryotic cells. However, the IME4 protein does not contain any previously identified nuclear localization segment (NLS) found in other genes; through this experiment we hope to be able to identify the portion of the protein that serves this function. Based on previous experiments in which segments of the IME4 gene were removed without interfering with nuclear localization, we hypothesize that the NLS is in the second half of the protein (C-terminal half). In order to test this idea, we attached the segment of IME4 that codes for the C-terminal half to a plasmid that encodes green fluorescent protein (GFP); because GFP does not undergo nuclear localization naturally, if GFP is found in the nucleus our hypothesis will be supported. To carry out this process, we first synthesized the modified copies of IME4 by amplification using the polymerase chain reaction (PCR). Then, we attached the IME4 gene to plasmids encoding GFP; these plasmids were then evaluated using agarose gel electrophoresis to determine whether they were formed correctly. Once the plasmids were confirmed, they were transformed into S. cerevisiae cells. Currently, the location of GFP is being examined using fluorescence microscopy.
Comments
2nd place, Poster