Functional Characterization of Novel lncRNA in Arabidopsis thaliana
Excluded from the Central Dogma, long non-coding RNA (lncRNA) are RNA of 200 nucleotides or more that lack the capacity to translate into protein. Following the discovery of lncRNA making up 99 percent of the human genome, there has been an effort to study the lncRNA of Arabidopsis thaliana, the model organism for plant science. Thirty years have passed since lncRNA were first characterized, but the roles of most lncRNA in A. thaliana remains unknown. Although studied lncRNA were generally found to play roles in regulating protein coding genes, particularly in developmental tissue, lncRNA should still be evaluated case-by-case to better understand their functions in this model organism. This project served to explore the role that lncRNA may play in regulating protein coding genes by characterizing the phenotypes of plants with mutations in a set of high-confidence lncRNAs identified as being either antisense or adjacent to protein coding genes through previous transcriptome analyses. This list of lncRNA also includes one lncRNA downstream of the FY gene, a controller of flowering time, to understand possible regulatory effects lncRNA may have on this significant stage of development. SALK and SAIL lines with TDNA insertions that targeted the lncRNA without disrupting the protein coding genes were ordered. For cases where the protein coding gene completely overlaps the antisense lncRNA, either antisense oligos were designed or the lncRNA were marked as candidates for CRISPR. Plants with the TDNA insertions were genotyped to confirm homozygous mutant status, and these plants were examined with initial phenotyping.
As a pre-med student, I was worried that I would have a difficult time understanding plant science or struggle with staying interested in plants for ten weeks. Wrapping up the program now, I am happy to say that both concerns proved to be false. Dr. Murphy and the rest of the supportive members of the Nelson lab continued to make my time at BTI constructive rather than degrading, and although we were tackling complicated topics that I have yet to encounter in school, I always had a better grasp of them before the end of the day. My main goal for this summer was to become more independent in the lab, and after just a couple of weeks, I felt comfortable extracting DNA, performing PCRs, and running gels, giving me the confidence and motivation to conduct more research in the future.