Pyramiding of cry toxins and methanol producing genes to increase insect resistance in cotton

Abdul Razzaq,Arfan Ali,Muhammad Mubashar Zafar,Aisha Nawaz,Deng Xiaoying,Li Pengtao,Ge Qun,Muhammad Ashraf,Maozhi Ren,Wankui Gong,Yuan Youlu

doi:10.1080/21645698.2021.1944013

Abstract

ABSTRACT The idea of enhanced methanol production from cell wall by pectin methyl esterase enzymes (PME) combined with expression of cry genes from Bacillus thuringiensis as a strategy to improve insect pest control in cotton is presented. We constructed a cassette containing two cry genes (cry1Fa and Cry32Aa) and two pme genes, one from Arabidopsis thaliana (AtPME), and other from Aspergillus. niger (AnPME) in pCAMBIA1301 plant expression vector using CAMV-35S promoter. This construction was transformed in Eagle-2 cotton variety by using shoot apex-cut Agrobacterium-mediated transformation. Expression of cry genes and pme genes was confirmed by qPCR. Methanol production was measured in control and in the cry and pme transformed plants showing methanol production only in transformed plants, in contrast to the non-transgenic cotton plants. Finally, insect bioassays performed with transgenic plants expressing cry and pme genes showed 100% mortality for Helicoverpa armigera (cotton bollworm) larvae, 70% mortality for Pectinophora gossypiella (pink bollworm) larvae and 95% mortality of Earias fabia, (spotted bollworm) larvae, that was higher than the transgenic plants expressing only cry genes that showed 84%, 49% and 79% mortality, respectively. These results demonstrate that Bt. cry-genes coupled with pme genes are an effective strategy to improve the control of different insect pests.

Highlights

Gossypium hirsutum L. is an important economical crop and one of the largest sources of natural fiber worldwide 1
These results indicated that Bacillus thuringiensis (Bt) cry-genes coupled with pme genes is a possible and useful strategy to control different insect pests and for lowering the resistance of insects against transgenic cotton varieties
Transgenic plants expressing simultaneously pme and cry genes were evaluated against different Lepidopteran insect pests and compared against non-transgenic and transgenic plants expressing only Bt cry-genes

Summary

Introduction

Gossypium hirsutum L. is an important economical crop and one of the largest sources of natural fiber worldwide 1. Bacillus thuringiensis (Bt) are soil bacteria, that produce different insecticidal proteins (named -endotoxins, such as Cry toxins), which have been successfully used against insect pests attack and several Cry insecticidal proteins have been transformed into cotton crops since 1996 4. The effectiveness of Bt -endotoxins started to decline due to the development of resistance by the insect pests 5. Despite of the proven substantial effects of transgenic Bt-cotton plants against insect attack, still is needed to improve this technology 6, for instance by combining with some enzymes involved in defense strategies against insect pests 7. The overproduction of enzymes, involved in insect defense, can be a good alternative to reduce the pest attack and development of insect resistance to Cry toxins 7

Methods

Results

Discussion

Conclusion