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A PCR detection method for mutations in receptor-protein genes from Busseola fusca potentially involved in Bt-resistance

Year: 2012
Type: dissertation
Abstract: Genetically modified (GM) crops attracted interest globally when use of these crops resulted in significant increases in yield and production.These increases were due to protection of crops from pests, weeds and diseases.However, evolution of resistance by pests threatens the continued efficacy of GM crops.One such example is the resistance to Cry1Ac toxin in Helicoverpa armigera (Lepidoptera: Noctuidae).Resistance in this pest was due to a mutation in the aminopeptidase N1 (APN) Cry receptor gene, encoding the receptor for Cry1Ac.Laboratory studies have indicated that species in families Noctuidae, Pyralidae and Plutellidae can develop resistance to Bttoxins.To date, field-evolved resistance has only been reported in Busseola fusca (Fuller) (Lepidoptera: Noctuidae) in South Africa, Helicoverpa zea (Boddie) (Lepidoptera: Noctuidae) in the south-eastern United States, Spodoptera frugiperda (J.E.Smith) (Lepidoptera: Noctuidae) in Puerto Rico, Pectinophora gossypiella (Saunders) (Lepidoptera: Gelechiidae) in India, Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) in northern China and Plutella xylostella (Linnaeus) (Lepidoptera: Plutellidae) in The Philippines and Hawaii.Resistance development in lepidopteran species is thus a common phenomenon.The stem borer B. fusca is a major insect pest to Bt-maize in the Vaalharts irrigation scheme (South Africa).The first official report of B. fusca resistance to Cry1Ab toxin was recorded in 2007, although farmers observed increased damage to Bt-maize from stem borers as early as 2004.A second report of resistance in an area nearby followed in 2009.No study has yet been done to determine the molecular mechanism of B. fusca resistance to Cry1Ab.As mentioned, a mutation in the APN receptor gene is responsible for H. armigera resistance to Cry1Ac.Although B. fusca has developed resistance to the B. thuringiensis Cry1Ab toxin, the binding-patterns and -sites of Cry1Ac and Cry1Ab are similar.Thus a similar mutation may be responsible for B. fusca resistance to Cry1Ab.Aminopeptidase, cadherin and alkaline phosphatase are the major Cry toxin receptors that have been identified in lepidopteran species.The present study was concerned with the investigation of mutations in these receptor genes.However, in order to study mutations, sequence data of receptor genes are essential.Degenerate primers were designed based on conserved regions observed in multiple protein sequence alignments of aminopeptidase N (isogenes 1 to 6), cadherin and alkaline phosphatase of several lepidopteran species.Primers were degenerate to take into consideration the Prof. C.C. Bezuidenhout, my supervisor.Thank you for all your advice and guidance with regard to this dissertation.I am privileged to have learnt so much from you and I thank you for all the opportunities that you have given me.Your encouragement, supervision and support have enabled me to develop an understanding of the subject.You motivated me when I did not get the results that I
Language: English
Open Access status: green
OpenAlex ID: W2181883979
Cites: 151
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