First report on CRISPR/Cas9 based in-vitro restriction of yellow gene in the oriental fruit fly, Bactrocera dorsalis (Hendel) (Tephritidae: Diptera)

Sangeeta Dash

doi:10.56093/ijas.v95i10.156193

Authors

Sangeeta Dash Ph.D. Research Scholar

https://doi.org/10.56093/ijas.v95i10.156193

Abstract

Bactrocera dorsalis is a serious pest that can cause significant financial losses. CRISPR/Cas9 is a reliable and effective approach for site-directed genome editing. The CRISPR/Cas9 system modifies DNA in a stable and heritable manner. Due to their complementary nature to the target gene, these artificially created single guide RNAs (sgRNAs) instruct the Cas9 endonuclease to produce double strand breaks in the gene sequence. This causes a functional disruption in target gene expression in the organism. The present investigation was, aimed at the validation of genetic targets for genetic biological control of this notorious pest via CRISPR/Cas9 mediated genome editing at ICAR- Indian Institute of Horticultural Research, Hessaraghatta, Bengaluru, 560089 on 2020-22. Gene yellow crucial for melanin pigmentation and body colouration in the insect was cloned followed by the synthesis of two (sg692 and sg915) off-target minimised guide RNA. The designed gRNAs were used for restriction assay to check the cleavage efficiency of sgRNAs. A successful in vitro cleavage of the gene was witnessed. Thus, our research proves the effectiveness of CRISPR/Cas9 mutagenesis as a suitable genetic control strategy for the management of pests of quarantine and economic significance.

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References

Andersen S O. 2012. Cuticular sclerotization and tanning. In Insect molecular biology and biochemistry. 167-192.

Arakane Y, Dittmer N T, Tomoyasu Y, Kramer K J, Muthukrishnan S, Beeman R W and Kanost M R. 2010. Identification, mRNA expression and functional analysis of several yellow family genes in Tribolium castaneum. Insect biochemistry and molecular biology. 40(3): 259-266.

Bai X, Zeng T, Ni X Y, Su H A, Huang J, Ye G Y, Lu Y Y, Qi Y X. 2019. CRISPR/Cas9‐mediated knockout of the eye pigmentation gene white leads to alterations in colour of head spots in the oriental fruit fly, Bactrocera dorsalis. Insect Molecular Biology. 28: 837–849.

Chen P, and Ye H. 2007. Population dynamics of Bactrocera dorsalis (Diptera: Tephritidae) and analysis of factors influencing populations in Baoshanba, Yunnan, China. Entomological Science. 10: 141-147.

Choo A, Crisp P, Saint R, O'Keefe L V and Baxter S W. 2018. CRISPR/Cas9‐mediated mutagenesis of the white gene in the tephritid pest Bactrocera tryoni. Journal of Applied Entomology. 142(1-2): 52-58.

Cong L, Ran F A, Cox D, Lin S, Barretto R, Habib N, Hsu P D, Wu X, Jiang W, Marraffini L A, and Zhang F. 2013. Multiplex genome engineering using CRISPR/Cas systems. Science. 339(6121): 819-823.

Deltcheva E, Chylinski K, Sharma C M, Gonzales K, Chao Y, Pirzada Z A, Eckert M R, Vogel J and Charpentier E. 2011. CRISPR RNA maturation by trans‐encoded small RNA and host factor RNase III. Nature. 471: 602–607.

Dong Z, Qin Q, Hu Z, Chen P, Huang L, Zhang X, Tian T, Lu C, and Pan M. 2019. Construction of a one‐vector multiplex CRISPR/Cas9 editing system to inhibit nucleopolyhedrovirus replication in silkworms. Virologica Sinica. 34: 444–453.

Heinze S D, Kohlbrenner T, Ippolito D, Meccariello A, Burger A, Mosimann C, Saccone G, and Bopp D. 2017. CRISPR-Cas9 targeted disruption of the yellow ortholog in the housefly identifies the brown body locus. Scientific reports. 7(1): 4582.

Li J, and Handler A M. 2019. CRISPR/Cas9-mediated gene editing in an exogenous transgene and an endogenous sex determination gene in the Caribbean fruit fly, Anastrepha suspensa. Gene. 691: 160-166.

Liu H, Zhang D, Xu Y, Wang L, Cheng D, Qi Y, Zeng L, Lu Y. 2019. Invasion, expansion, and control of Bactrocera dorsalis (Hendel) in China. Journal of Integrative Agriculture. 18: 771–787.

Meccariello A, Tsoumani K T, Gravina A, Primo P, Buonanno M, Mathiopoulos K D, and Saccone G. 2020. Targeted somatic mutagenesis through CRISPR/Cas9 ribonucleoprotein complexes in the olive fruit fly, Bactrocera oleae. Archives of Insect Biochemistry and Physiology. 104(2): e21667.

Meccariello A, Monti S M, Romanelli A, Colonna R, Primo P, Inghilterra M G, Del Corsano G, Ramaglia A, Iazzetti G, Chiarore A, and Patti F. 2017. Highly efficient DNA-free gene disruption in the agricultural pest Ceratitis capitata by CRISPR-Cas9 ribonucleoprotein complexes. Scientific reports. 7(1): 10061.

Meng L W, Yuan G R, Lu X P, Jing T X, Zheng L S, Yong H X, Wang J J. 2019. Two delta class glutathione S-transferases involved in the detoxification of malathion in Bactrocera dorsalis (Hendel). Pest Management Science. 75: 1527–1538.

Montague T G, Cruz J M, Gagnon J A, Church G M and Valen E. 2014. CHOPCHOP: a CRISPR/Cas9 and TALEN web tool for genome editing. Nucleic Acids Research. 42: 401-407.

Moon T T, Maliha I J, Khan A A M, Chakraborty M, Uddin M S, Amin M R and Islam T. 2022. CRISPR‐Cas genome editing for insect pest stress management in crop plants. Stresses. 2: 493–514.

Nguyen T N, Mendez V, Ward C, Crisp P, Papanicolaou A, Choo A, Taylor P W and Baxter S W. 2021. Disruption of duplicated yellow genes in Bactrocera tryoni modifies pigmentation colouration and impacts behaviour. Journal of Pest Science. 94: 917-932.

Paulo D F, Cha A Y, Kauwe A N, Curbelo K, Corpuz R L, Simmonds T J, Sim S B and Geib S M. 2022. A unified protocol for CRISPR/ Cas9‐mediated gene knockout in tephritid fruit flies led to the recreation of white eye and white puparium phenotypes in the melon fly. Journal of Economic Entomology. 115: 2110–2115.

Pradhan S K, Karuppannasamy A, Sujatha P M, Nagaraja B C, Narayanappa A C, Chalapathi P, Dhawane Y, Bynakal S, Riegler M, Maligeppagol M and Ramasamy A. 2023. Embryonic microinjection of ribonucleoprotein complex (Cas9+ sgRNA) of white gene in melon fly, Zeugodacus cucurbitae (Coquillett)(Diptera: Tephritidae) produced white eye phenotype. Archives of Insect Biochemistry and Physiology. 114(4): e22059.

Rajan V V, Kumar H, Parvathy M, Bhargava C, Ashok K, Pradan S K, ANU C, Aravintharaj R, and Asokan R. 2023. Modifying oviposition behaviour of the Oriental fruit fly, Bactrocera dorsalis (Hendel) to obtain uniform G0. Pest Management in Horticultural Ecosystems. 29(1).

Sim S B, Kauwe A N, Ruano R E, Rendon P, and Geib S M. 2019. The ABCs of CRISPR in Tephritidae: developing methods for inducing heritable mutations in the genera Anastrepha, Bactrocera and Ceratitis. Insect Molecular Biology. 28(2): 277-289.

Wan X, Liu Y, and Zhang B. 2012. Invasion history of the oriental fruit fly, Bactrocera dorsalis, in the Pacific-Asia region: two main invasion routes. PLoS One. 7: 36176.

Wang J, Wang H, Liu S, Liu L, Tay W T, Walsh T K, Yang Y, and Wu Y. 2017. CRISPR/Cas9 mediated genome editing of Helicoverpa armigera with mutations of an ABC transporter gene HaABCA2 confers resistance to Bacillus thuringiensis Cry2A toxins. Insect biochemistry and molecular biology. 87: 147-153.

Wittkopp P J, True J R, Carroll S B. 2002. Reciprocal functions of the Drosophila yellow and ebony proteins in the development and evolution of pigment patterns. Development. 129: 1849–1858.

Zhang L and Reed R D. 2017. A practical guide to CRISPR/Cas9 genome editing in Lepidoptera. Diversity and Evolution of Butterfly Wing Patterns: An Integrative Approach. 24: 155–172.

Zheng W, Li Q, Sun H, Ali MW and Zhang H. 2019. Clustered regularly interspaced short palindromic repeats (CRISPR)/ CRISPR‐associated 9‐mediated mutagenesis of the multiple edematous wings gene induces muscle weakness and flightlessness in Bactrocera dorsalis (Diptera: Tephritidae). Insect Molecular Biology. 28: 222–234.

Zhu Y, Qi F, Tan X, Zhang T, Teng Z, Fan Y, Wan F, Zhou H. 2022. Use of Age-Stage, Two-Sex Life Table to Compare the Fitness of Bactrocera dorsalis (Diptera: Tephritidae) on Northern and Southern Host Fruits in China. Insects. 13: 258.