Oligonucleotides applied in genomics, bioinformatics and development of molecular markers for rice and barley

• Main Author: Liu, Shaolin, 1968-
• Format: Others
• Language: en
• Published: McGill University 2004
• Subjects: Oligonucleotides.; Genetic markers.; Rice > Molecular genetics.; Barley > Molecular genetics.
• Online Access: http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=85569

A genome sequence can be conceptualized as a 'book' written with four nucleotide 'letters' in oligonucleotide (oligo) 'words'. These words can be used in genomics, bioinformatics and the development of molecular markers. The whole-genome sequence for rice (Oryza sativa L.) is almost finished and has been assembled into pseudomolecules. For barley ( Hordeum vulgare L.) expressed sequence tags (ESTs) have been assembled into 21,981 tentative consensus sequences (TCs). The availability of such sequence information provides opportunities to investigate oligo usage within and between genomes. For the first of three studies reported in this thesis, a C++ program was written to automatically design oligos that are conserved between two sets of sequence information. In silico mapping between rice coding sequences (CDS) and barley TCs indicated that oligos between 18 and 24 bp provide good specificity and sensitivity (83% and 86%, respectively, for 20mers). Conserved oligos used as PCR primers had a high (91%) success rate on barley lines. Sequencing of PCR products revealed conservation in exon sequence, size and order between barley and rice. Introns were not conserved in sequence but were relatively stable in size. Map locations of eight new markers in barley revealed both genome colinearity and rearrangements between barley and rice. The second study reported in this thesis examined word frequency within the rice genome. A non-random landscape composed of high-frequency and low-frequency zones was observed. Interestingly, high-frequency words seemed to be rice specific while single-copy words were gene specific and conserved across species. As in the first study, oligos of 12 bp or less were not specific, and 18 bp seemed to be a critical length for the specificity of oligos. The third study reported in this thesis involved the development of molecular markers for known genes using public sequence information. Six new polymorphic markers were d