<p> Until a little over a decade ago, the regulatory roles of small RNAs (sRNAs) in
prokaryotes were largely undetected. Since then, there has been an explosion in the discovery of
novel sRNA sequences and we have begun to understand their functions and mechanisms of
regulation. The identification and characterization of sRNAs from different organisms have
largely been achieved through computational and experimental approaches that focus on
sequence elements in intergenic regions. Based on these previously established techniques, we
have developed and applied a new bioinformatics approach to search for highly conserved
sequences in unannotated intergenic regions from several bacterial genomes, which may contain
new sRNA sequences. Through this search, we have identified seven candidate sequences that are conserved at the primary sequence level, and some of the secondary structure motifs are also conserved among multiple bacteria genomes. When we examined those seven candidates
experimentally, it was found that when the expression of one mutated candidate (rUIG0803 _ 4D) was induced at the RNA level, minor morphological changes and a delayed lethal phenotype was elicited. The expression of the RNA also may result in the altered expression of kanamycin
kinase and glycerol kinase, as indicated by the mass spectrometry data. Experimental
characterizations of eight previously identified sRNAs from literature with functions unknown
have also been performed but no apparent phenotypic phenomenon was observed in this project,
which indicated that all or some of those 8 sRNAs might not play any regulatory roles in cells, or
their roles need to be characterized through other genetic screens. To further search for RNA
sequences with regulatory functions, we created a library of random DNA transcript using the
Lambda Phage genomic DNA. Preliminary screening efforts show that three of the 192 clones
screened could trigger reduced cell growth when their RNA was overexpressed. This study marks
the first use of a bioinformatics approach that uses primary sequence and secondary structure
information to search for sRNAs in the unannotated intergenic region. Moreover it also marks the first time that the effects of introducing random lambda phage RNA in an E. coli host. </p> / Thesis / Master of Science (MSc)
| Identifer | oai:union.ndltd.org:mcmaster.ca/oai:macsphere.mcmaster.ca:11375/21611 |
| Date | 05 1900 |
| Creators | Zhu, Rebecca |
| Contributors | Li, Yingfu, Biochemistry and Biomedical Sciences |
| Source Sets | McMaster University |
| Language | English |
| Detected Language | English |
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