| Hexaploid oat (Avena sativa L.) is an annual herbaceous plant in the Poaceae, which is cultivated worldwide for food and fodder, as well as for specialty uses in diet therapy and nutrition.Genetic resources of oat are abundant and variable. Oats that thresh free from their hulls (naked oats) are cultivated widely in China, while hulled oats are most commonly grown in the other countries. Oats are generally considered as a healthy food because they are high in fibre, beneficial oils, and good-quality protein. Oat protein is nearly equivalent in quality to soy protein, which World Health Organization research has shown to be nutritionally equivalent to meat, milk, and egg protein.Tools for genetic improvement of oat are being developed. However, genomic analysis of oat is challenging due to its large, repetitive hexaploid genome. Studies reported of this thesis were conducted to better understand the cytogenetic organization of the oat genome, and to provide the oat research community with new tools for characterizing genetic variability in oat, and the application of genomic tools to discover genetic factors affecting flowering time, an important adaptive characteristic of oat. In this study, we have sequenced and analysed18D-specific library from hexaploid oat(Avena sativa L.), then chose some special sequences as FISH probes to detect species in Avena; we also investigated response to vernalization and photoperiod in a population of384diverse oat varieties (Integrated Oat Improvement, IOI). We then cloned and sequenced the fragments of a candidate gene and analysed the genetic diversity of storage proteins in oat. The main results are as follows:(1) We have sequenced, assembled, and characterized a set of complexity-reduced genomic clones derived from a chromosome18D-specific library from hexaploid oat. Sequences from314clones were assembled into99contigs of identical or nearly identical sequence. The Censor tool was used to identify similarity to known and characterized repeat sequences in RepBase. Eight repeat classes were scattered throughout50contigs, with most repeats belonging to seven transposon and retrotransposon classes. After accounting for known repeats, additional matches to orthologous genes from other species were identified in24regions of22contigs, and an additional47regions matched genomic sequences from oat and other related species. These results provide information about the types and density of transposable elements in the oat genome, as well as the potential for identifying unique or chromosome-specific sequence elements in oat. Overall, these results predict a low success rate in identifying chromosome-specific coding regions in oat through chromosome isolation and genome complexity reduction.(2) The probes of A336and pAml were used to label chromosomes in mitotic metaphase plates of A. sativa (AACCDD). The results are as follows:a) chromosomes in A. sativa were labelled in the centromere by probe A336. b) A genome and D genome are prior to be labelled by probe A336. c) all chromosomes would be labelled with the high concentration of probe. These results suggested probe A336may be used to label centromeres in A. sativa.(3) The probes of A250, A305, and A436were used to label chromosomes in mitotic metaphase plates of A. canariensis (AcAc). As a result, all chromosomes in A. canariensis probes were labelled separately by three probes A250, A305, and A436. This result indicated there might be a close relationship bettween18D chromosome and Ac genome.(4) The probes of pITS and pAml were used to label chromosomes in mitotic metaphase plates of A. sativa (AACCDD) and A. canariensis (AcAc). The results are as follows:a) chromosomes in C genome from A. sativa were labelled by probe pAm1. b) six telomeres in A/D genome from A. sativa were labelled by probe pITS. c) four telomeres in A. canariensis (Ac) were labelled by probe pITS. Together with previous results that two telomeres in A genome and four in D genome from A. sativa were labelled by probe pITS, we could infer there might be a similar distribution of pITS bettween Ac genome and D genome.(5) The probes of A3-19, pAs120a and pAml were used to label chromosomes in mitotic metaphase plates of A. sativa (AACCDD), A. maroccana (AACC), A. vaviloviana (AABB), A. longiglumis (A1A1), and A. canariensis (AcAc). The main results are as follows:a) all signals labelled by A3-19are detected in telomeres or subtelometres. b) two telomeres and two subtelomeres in A genome were labelled by probe A3-19. c) similarly, two telomeres and two subtelomeres in B genome were labelled by probe A3-19. d) two subtelomeres in Ac genome were labelled by probe A3-19. e) two subtelomeres in D genome were labelled by probe A3-19. These results suggested the follows:a) distribution of A3-19on Ac genome is different to A1genome, but similar to D genome. b) A genome is close to B genome.(6) The IOI population was at different controlled temperatures and light regimes to determine the time to flowering under each condition. It was found that day-length had a greater influence on heading date, and that most oat varieties did not flower under short- day conditions. Based on the heading date, winter types and spring types were identified. Taking into account of geographical origin, the results further demonstrated that vernalization insensitive forms might occur in the north Mediterranean region and the adjacent northern territories.(7) Six fragments of vernalization and photoperiod candidate genes were used to analyse genetic diversity between winter type and spring type. The fragment sequences were clustered into spring and winter groups by TCS network analysis. Dancer_CDC, Kanota, Ogle, Starter, Buffalo, and Heinrich were contained in spring group of vernalization gene fragments (VRN1-6, VRN1-19) and photoperiod gene fragments (VRN3-23, VRN3-24), while the Kingfisher, Fleuron, Wistar, Bage1419/36, and Landhafer were include in winter group. However, the winter group also included Kanota VRN1-1, as well as Kanota and Ogle VRN1-18. These results indicated that there should be some genetic difference between winter type and spring type in oat population. These sequences provided a guide to explore the vernalization and photoperiod genes in other temperate cereals.(8) SDS-PAGE was used to analyse the genetic diversity of globulin, glutelin, prolamin between winter type and spring type in IOI population. More than seven protein bands were found in each accession. Globulin was focused on the region of50KD, whereas glutelin was manily distributed in the region between26KD and33KD. However, prolamin was scattered in the region less than60KD. There is a low polymorphism for globulin, and relatively high in glutelin, prolamin bands are most dispersed. Each accession displayed a variable distribution of bands. However, there was a homogeneous distribution of prolamin bands among spring types, separately. These results indicated the difference of spring type and winter type cannot be clarified well by their seeds storage proteins diversity.As an overall summary:probes of A250, A305, and A436suggested there might be a close relationship bettween18D chromosome and Ac genome; probe of pITS and A3-19showed there might be a similar distribution of pITS bettween Ac genome and D genome. These results indicated Ac genome is close to D genome, and A. canariensis (AcAc) could be the D genome donor of Avena sativa. Vernalization and photoperiod had a great influence on heading date of spring type oat and winter type in the IOI population; there should be some genetic difference between winter type and spring type in oat population; there is a relative abundance of genetic diversity of protein between winter type and spring type in IOI population but winter type and spring type cannot be clarified well by their storage protein diversity. These results indicated there are differences in vernalization and photoperiod response that are possibly attributable to specific candidate genes segregating in the IOI population. |
PDF Full Text Request