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2.22 analysis (Edwards et al., 1991 ). The extradion procedure uses ionic detegents. such as SDS or CTAB. which should be removed by phenol extraction because they inhibit Taq polymerase activity. Residual traces of phenol, which also inhibits PCR. should be removed by chlorofom : isoamyl alcohol(24:l) extraction or by ethanol prdpitation of the DNA (Newton and Graham. 1994). Concems have been expressed regarding the reliability of RAPD markers. These concems are based on the fact that random primers do not bind 100 % to target DNA. as is the case with primers based on specific sequence information in other PCR assays. The accuracy of random primers in RAPD analysis compared to RFLP analysis was tested in the detedion of geneüc relationships within and among cruciferous species (Thomann et al., 1994). They found a discrepancy for interspecific but not intraspecific relationships between RFLP and WPD data due to the false scoring of non-homologous amplified sequences as homologous. DNA extraction procedures can affect repeatability of RAPD markers as well. It is important to use consistent methods to optimize the template concentration relative to the primer to minimize cornpetition for primer sites. Themocyclers have been tested for reliability and different machines have produced repeatable RAPD products (Smith and Chin, 1992; Weeden et al., 1992; Mailer et al.. 1994). Vanous concentrations of Taq polymerase and numbers of cycles of PCR also produce repeatable RAPD products (Smith and Chin. 1992). Methods of scoring RAPD products may also result in non-repeatable results. Faint bands tend to be less repeatable and therefore should not be scored as RAPD pmducts unless tested repeatedly. The optimization and
2.23 maintenance of DNA extradion procedures and PCR conditions should minimue concems about repeatability of RAPD markers. The advantages of using RAPD markers have ben shown in the identification of useful genetic markers and esümating genetic relationships or diversity in several crop species. RAPD markers are dominant and therefore deted less polyrnorphism per locus than RFLPs. only two versus multiple alleles in RFLP. but they also do not require as much DNA and time as RFLP assays. This has led to suggestions that RAPD analysis be used in mapping cultivar genomes. such as rice. where the wide hybridizations required to produce enough polymorphisrn for RFLP analysis often result in sterility or poor growth (Mackill, 1995). Williams et al. (1990) also showed that RAPD markers could saturate the soybean map by flling in areas not detected with RFLP analysis. RAPD marken linked to specific traits can serve as an alternative to morphological markers, allowing eariier selection for disease resistance in rice varieties (Naqvi, et al., 1995) and for outcrossed tetraploid alfalfa plants (Gjuric and Smith. 1996). RAPD markers have also been used to distinguish genetic relationships among cultivars in B. napus (Mailer et al., 1994). pea (Bagheri et al.. 1995) and potato (Demeke et al., 1996) and between species in Brassica (Demeke et al., 1992) . Similar genetic relationships using RAPD and RFLP analysis have been reported in B. napus breeding lines (Hallden et al.. 1994) and B. oleracea genotypes (dos Santos et al., 1994). These studies came after reports that RAPD markers were not as reliable as RFLP in detecting genetic relationships between cniciferous species (Thormann et al.,
Page 1 and 2: The Application of Ooubled Haploid
Page 3 and 4: COPYRIGET P1ERbIISSION PAGE A Thesu
Page 5 and 6: 111 application in 8. rapa cultivar
Page 7 and 8: v TABLE OF CONTENTS ABSTWT ACKNOWLE
Page 9 and 10: VI1 LIST OF TABLES TABLE Page EFI',
Page 11 and 12: lx LlST OF FIGURES FIGURE Page Prod
Page 13 and 14: 1.2 performance. Production of DH p
Page 15 and 16: History of Canola Produdion of Bras
Page 17 and 18: Economic Importance and Distributio
Page 19 and 20: 2.5 genes involved in recognizhg an
Page 21 and 22: 2.7 There is potential for loss of
Page 23 and 24: 2.9 their stability under different
Page 25 and 26: 2.1 1 selected and selfed to create
Page 27 and 28: 2.1 3 production (Mathias and Wbbel
Page 29 and 30: 2.1 5 traits have low heritability.
Page 31 and 32: 2.17 opposite strands of template D
Page 33 and 34: Figure 2.3. The AFLP procedure usin
Page 35: 2.21 non-specifïc arüfacts which
Page 39 and 40: Cornparison of Bud Pollination and
Page 41 and 42: INTRODUCTION Production of doubled
Page 43 and 44: Effkiency lndex = total number of s
Page 46 and 47: Application of doubfeâ haploid dev
Page 48 and 49: Brassita rapa represents approximat
Page 50 and 51: 4.5 and a daylnight temperature of
Page 52 and 53: 4-7 plants within each tent using a
Page 54 and 55: 4.9 Days to Flowanng Reward OH Iine
Page 56 and 57: 4-11 In Canan, Reward Cja and Cl,,
Page 58 and 59: 4.13 counterparts. The extra genera
Page 60 and 61: 4-15 composite population can eithe
Page 62: Table 4.2. Mean parameter values fo
Page 68 and 69: Table 4.8. Correlation of days to f
Page 72 and 73: Detedion of Genetic Variation in Bm
Page 74: INTRODUCTION The production of doub
Page 77 and 78: 5-6 represented one sample from C,
Page 79 and 80: 5-8 the percent of polymorphism pre
Page 81 and 82: 5-10 and C, and a similar level of
Page 83 and 84: Table 5.2. Polymorphic levels in Cl
Page 86 and 87:
5-15 Table 5.5. Proportion of polyr
Page 88 and 89:
6. GENERAL DlSCUSSlON AND CONCLUSIO
Page 90 and 91:
6-3 B. rapa OH Iines. Seleded DH Ii
Page 92 and 93:
Abramson, RD. 1995. Thermostable DN
Page 94 and 95:
Diers, B.W. and Osbom, T.C. 1994. G
Page 96 and 97:
Jasieniuk, M., Brill&Babel, AL. and
Page 98 and 99:
Narasimhulu, SBw and Chopra, U.L. 1
Page 100 and 101:
Schuler, T.J., Hutcheson, O.S. and
Page 102 and 103:
Williams, J.G.K., HanMey, M.K., Raf
Page 104 and 105:
Appendix 12. Data matrix of RAPO pr
Page 106 and 107:
Apperidi 1. (-nueû) Month D~Y Temp
Page 108 and 109:
Mean Max [mm)
Page 110 and 111:
2 3 4 5 6 7 8 9 10 11 12 13 14 15 1
Page 112:
Append'bc 4. FM measummmts fw param
Page 116 and 117:
Locaüon Plant Id. RedTcate €MG D
Page 118 and 119:
C24 C24 C2-4 C2-8 C2-8 C2-8 C2-8 c2
Page 120 and 121:
Location Plant Id. Replicate €MG
Page 122 and 123:
Locaüori Plant ld. Replicate EMG O
Page 124 and 125:
Appendix 10. The mean performance o
Page 127:
f i . I 1 t a I 1 I I t t c i r' !m
Page 130 and 131:
Appendii 12. Data matrix of RAPO pm
Page 133:
Appendix 13. @mtïnued) Primer Comp
Page 138:
Appendk 14. (continuedl Primer Comp
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