The use of microsatellite markers has matured and become commonplace for
plant genome analyses and is now poised for widespread practical application
in sugarcane. Sequence Tagged Microsatellite Site (STMS) amplification is the
most prevalent microsatellite-based approach and involves the amplification of
a microsatellite by designing primers that flank and hence define the
microsatellite site, revealing variation in the length of repeat motifs between
individuals. Twenty-six microsatellite primer pairs received from the
International Sugarcane Microsatellite Consortium (ISMC) were evaluated and
the STMS protocol was optimised to ensure robust and reproducible results.
The objectives of this study were to use STMS for sugarcane parentage
analysis and fingerprinting. Previously, Restriction Fragment Length
Polymorphism (RFLP) marker data had suggested that the parentage of a
genetic mapping population, sugarcane cross AA40 (N18 x CP57/614), was
incorrect. Based on the assertion that the incorrect parentage was as a result of
either mislabelling at planting or at seed collection, microsatellite parentage
analysis was carried out on eight potential parent pairs (13 cultivars). A total of
75 markers were scored with non-parental bands (12 on average) being
observed for all of the potential parent pairs and none could be identified as the
true AA40 parents. It has been suggested in other plant species that PCR
artefacts could give rise to non-parental bands and to investigate this the
marker data of single parent DNA reactions and pooled parent pair DNA
reactions or 'synthetic offspring' were compared. The results suggested that
either a certain percentage of non-parental bands, perhaps 10% (maximum
value observed), should be tolerated in microsatellite parentage analysis or a
marker should only be considered to be discriminating for parentage if it is
absent in both the parents and the pooled parent pair amplifications.
Fingerprinting of 20 cultivars using 14 microsatellite primer pairs was conducted
to evaluate the potential of the STMS approach for sugarcane varietal
identification. It was found that only two microsatellite primer pairs were
required to discriminate between all 20 cultivars with a theoretical number of
non-differentiated pairs of cultivars (XK) of only 0.03. This estimator was used to
determine the approximate number of microsatellites necessary for large-scale
sugarcane fingerprinting. / Thesis (M.Sc.)-University of Natal, Durban, 2002.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:ukzn/oai:http://researchspace.ukzn.ac.za:10413/4687 |
| Date | January 2002 |
| Creators | Hack, Simon Matthew. |
| Contributors | Huckett, Barbara I., Butterfield, Mike. |
| Source Sets | South African National ETD Portal |
| Language | English |
| Detected Language | English |
| Type | Thesis |
Page generated in 0.0025 seconds