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Isolate IG1 Incidence, distribution and characteristics of major tomato leaf curl and mosaic virus diseases Isolate IG1 tested positive in PCR with primers PAL1v1978 and PAR1c715 (Figure 3.12 C). It was also positive when primers PARAv 494 and PARAc 1048 were used. Purified sample IG1 DNA was cloned with a TOPO vector, and when sequenced gave a forward viral sense sequence of 482 nt in size, and a reverse coat protein sequence of 521 nt. The coat protein was part of the upper part of the circular DNA presented in Figure 3.13. From these partial sequences, primers PTLCV-UGrep2r, PTLCV-UGcp1f and PTLCV- UGcp1r were synthesized (Table 3.12). Isolate RL5 (DNA Intergenic Region) In PCR, isolate RL5 reacted positively to TYLCV-Is specific primers (PTYCRv 21 and PTYCRc 287) as indicated in Figure 3.12E, and was also positive to TYLCV-Is in an experiment with primers PTYC2v 1499 and PTYAL1c 2196 (Figure 3.13F and Table 3.13). Purified DNA (fluorimeter reading of 268 ng/µl) of type isolate RL5 amplified with primers PTYCRv 21 and PTYIRc 287 for the TYLCV-Is common region auto produced a 277 nt TYLCV sequence (excludes vector and primers segments as well as overlapping) (Figure 3.14). From these results, isolate RL5 was positive to TYLCV-Is, and was therefore tentatively named TYLCV-Ugandan isolate (TYLCV-UG). 82
Incidence, distribution and characteristics of major tomato leaf curl and mosaic virus diseases Table 3.9: Reaction of squashed and dot-blotted tomato geminivirus samples to the general, specific, and mixed probes (See figure 3.11 A, C, D, and F) Sample Over night exposure 2 Days Exposure Code General probe Mixed probe General probe Specific probe BGMV BGMV + TYLCV-EG BGMV TYLCV-EG RL1 0³ +¹ 0 ++ MB7 0 RL2 0 MB9 0 MB2 -² K3 0 RL5 - MB6 0 IG2 - IG1 ++ IG7 + IG10 + MB8 0 ISO30 +++ ISOPOT 0 TYLCV +++ BGMV +++ Healthy - + 0 + ++ 0 ++ + 0 + + - + ++ 0 +++¹ + - ++ - 0 + - - + + ++ ++ 0 + 0 0 + 0 + 0 ++ 0 +++ 0 + 0 ++ +++ +++ +++ - +++ - - - - ¹ (+, …, +++) intensity of positive reaction; ²(-) negative reaction; ³ (0) means that the sample was not tested with that particular probe. 83
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Incidence, Distribution and Charact
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Charles SSEKYEWA Incidence, Distrib
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CONTENTS 1 CHAPTER 1...............
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ACKNOWLEDGEMENT The following perso
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vii 7 mosaic virus (CMV), Alfalfa m
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ix 9 Consequently, our findings con
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SAMENVATTING Tomaat is een economis
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xiii 13 Beide genhomologie percenta
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xv 15 dit onderzoek geven ook aan d
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LIST OF TABLES xvii 17 Table Page 1
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ACRONYMS AND ABBREVIATIONS xix 19 A
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Abbreviation Word in full PCR Polym
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Incidence, distribution and charact
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Table 2.1: The orthography of some
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8 ANNEXES Incidence, distribution a
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Table A.1.1 continued Incidence, di
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Annex 5: Incidence, distribution an
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