THERMODYNAMIC STABILITY OF MRNAS IN PLANTS USING MFOLD: ENHANCED STABILITY OF THE TORTULA RURALIS ADLH7B6 TRANSCRIPT

AN ABSTRACT OF THE THESIS OF William Charles Phillips, for the Master of Science degree in Plant Biology, presented on April 7, 2008, at Southern Illinois University Carbondale. TITLE: THERMODYNAMIC STABILITY OF MRNAS IN PLANTS USING MFOLD: ENHANCED STABILITY OF THE TORTULA RURALIS ADLH7B6 TRANSCRIPT MAJOR PROFESSOR: Dr. Wood The cDNA of animals, angiosperms and T. ruralis contains ALDH7, which is an enzyme from a family that is stress responsive (Wood and Krayesky 2002). The ALDH7 enzyme family is characterized into two different groups called the A and B sub-family of enzymes. They are either: animal specific and plant specific with the B sub-family being plant specific. The ALDH7 ortholog (ALDH7B6) is from the desiccation-tolerant moss T. ruralis. This homolog happens to be highly expressed in control tissues, and steady-state mRNA transcript abundances are unchanged during different types of stress induction of mRNA (Wood et al 2002). To sufficiently investigate the stability of the ALDH7B6 nucleotide sequence of T. ruralis, a variety of search tools we used. Similar sequences to the ALDH7B6 are available on the web through the use of the BLAST search engine, which can be found at http://www.ncbi.nlm.nih.gov/. Sequences are selected based on their E values and PAM scores which is percent accepted scoring matrix. The selected orthologs are entered into bioinformatics tools to complete the production of tables to express the similarity among the sequences. The selected program for the similarity among sequences is the Lalign program. The Lalign program is located at this website: http://www.ch.embnet.org/software/LALIGN_form.htm. For the actual measurement of the mRNAs stability, the Mfold program is available. Mfold is located at the following web address: http://www.bioinfo.rpi.edu/applications/mfold/old/rna/. This program produces a selection of the top most optimal foldings of the mRNA\u27s secondary structure. Included is a Delta G score, which describes the thermodynamics of the structure. From these programs, data are interpreted, and statistical analyses are preformed. The main important finding is that T. ruralis has the most negative Delta G for its complete nucleotide sequence (-734.4 kcal/mol), and its Delta G is significant based on it being more than two z scores away from the mean of the complete nucleotide sequence Delta G scores. This means that the secondary structure contains the most negative free energy, so it requires the most energy to change its folding. Also, T. ruralis has the most negative nucleotide coding region with a Delta G score of (-568.1 kcal/mol), yet this score isn\u27t significant based on two z scores away from the mean. The Delta G scores for the conserved regions are similar, which is expected based on the sequences being conserved and of similar lengths. Tortula ruralis has the most negative 3\u27 UTR when computed by kcal/mol/base. When describing the data this way, animals have a more negative 5\u27 UTR than plants, with the exception of O. sativa. The GC content is above 40% for all species for their complete nucleotide sequence and nucleotide coding regions. The GC content is an important contributor to the results from Mfold. Along with the thermodynamic scores, analyze were; the most optimal folding of the 5\u27 UTR; complete nucleotide sequence; nucleotide coding region; and the 3\u27 UTR for all of the species, (except the 3\u27 UTR and 5\u27 UTR of B. napus ALDH7 because it was composed of gDNA). Summarized and compared these nucleotide sequences and conclusions were based on similarities. Conclusions: The overall sequence length and GC content of a nucleotide sequence determine the stability as described by Mfold