Please wait a minute...
Journal of Integrative Agriculture  2022, Vol. 21 Issue (10): 2818-2832    DOI: 10.1016/j.jia.2022.07.033
Crop Science Advanced Online Publication | Current Issue | Archive | Adv Search |
Expression profiling of transgenes (Cry1Ac and Cry2A) in cotton genotypes under different genetic backgrounds

Kashif NOOR1, Hafiza Masooma Naseer CHEEMA1, Asif Ali KHAN2, Rao Sohail Ahmad KHAN3

1 Department of Plant Breeding and Genetics, University of Agriculture, Faisalabad 38040, Pakistan

2 Institute of Plant Breeding and Biotechnology, Muhammad Nawaz Shareef University of Agriculture, Multan 60000, Pakistan

3 Center of Agricultural Biochemistry and Biotechnology, University of Agriculture, Faisalabad 38040, Pakistan

Download:  PDF in ScienceDirect  
Export:  BibTeX | EndNote (RIS)      
Abstract  

Transgenic cotton carrying the Cry1Ac gene has revolutionized insect pest control since its adoption, although the development of resistance in insect pests has reduced its efficacy.  After 10 years of cultivating Bacillus thuringiensis (Bt) cotton with a single Cry1Ac gene, growers are on the verge of adopting Bt cotton that carries the double gene (Cry1Ac+Cry2A) due to its better effectiveness against insect pests.  Thus, the current study was designed to evaluate the role of each gene in the effectiveness of Bt cotton carrying the double gene.  The expression levels of the Cry1Ac and Cry2A genes were evaluated in the leaves of 10 genotypes (2 parents and 8 F1 hybrids) at 30 days after sowing (DAS), while samples of leaves, bolls and flowers were taken from the upper and lower canopies at 70 and 110 DAS.  The F1 hybrids were developed through reciprocal crosses between two Bt (CKC-1, CKC-2) and two non-Bt (MNH-786, FH-942) parents.  The differential expression of transgenes was evaluated through Enzyme Linked Immuno-Sorbent Assay (ELISA).  The results showed that the MNH786×CKC-1 hybrid had the highest concentrations of Cry1Ac gene at 30 DAS (3.08 µg g–1) and 110 DAS (1.01 µg g–1) in leaves.  In contrast, the CKC-2×MNH-786 hybrid showed the lowest concentrations of Cry1Ac gene at 30 DAS (2.30 µg g–1) and 110 DAS (0.86 µg g–1).  The F1 hybrid FH-942×CKC-2 showed the highest concentrations of Cry2A gene at 30 DAS (8.39 µg g–1) and 110 DAS (7.74 µg g–1) in leaves, while the CKC-1×MNH-786 hybrid expressed the lowest concentrations of Cry2A gene at 30 DAS (7.10 µg g–1) and 110 DAS (8.31 µg g–1).  A comparison between the two stages of plant growth showed that leaves had the highest concentrations at 30 DAS, whereas the lowest concentrations were observed at 110 DAS for both genes in leaves.  When the expression pattern was compared between various plant parts in genotype CKC-2, it was found that leaves had higher concentrations of Cry1Ac (3.12 µg g–1) and Cry2A (8.31 µg g–1) at 70 DAS, followed by bolls (Cry1Ac (1.66 µg g–1) and Cry2A (8.15 µg g–1)) and flowers (Cry1Ac (1.07 µg g–1) and Cry2A (7.99 µg g–1)).  The genotype CKC-2 had higher concentrations of Cry1Ac (3.12 µg g–1) and Cry2A (8.31 µg g1) in the upper canopy but less accumulation (2.66 µg g–1 of Cry1Ac, 8.09 µg g–1 of Cry2A) in the lower canopy at 70 DAS.  Similarly, at 110 DAS, the expression levels of Cry1Ac and Cry2A in upper and lower canopy leaves were 1.52 and 7.92 µg g–1, and 0.99 and 7.54 µg g–1, respectively.  Hence, the current study demonstrates that different genotypes showed variable expression for both of the Cry1Ac and Cry2A genes during plant growth due to different genetic backgrounds.  The Cry2A gene had three-fold higher expression than Cry1Ac with significant differences in expression in different plant parts.  The findings of this study will be helpful for breeding insect-resistant double-gene genotypes with better gene expression levels of Cry1Ac and Cry2A for sustainable cotton production worldwide.

Keywords:  transgenic cotton breeding        transgene expression        double gene        insect resistance  
Received: 08 March 2021   Accepted: 08 July 2021
Fund: The authors are further thankful to Higher Education Commission, Pakistan for providing funds, Center of Excellence in Molecular Biology, Lahore and University of Agriculture, Faisalabad, Pakistan for their resources, support and encouragement in this study.  
About author:  Kashif NOOR, E-mail: kashifnoor48@yahoo.com; Correspondence Hafiza Masooma Naseer CHEEMA, E-mail: masooma@uaf.edu.pk

Cite this article: 

Kashif NOOR, Hafiza Masooma Naseer CHEEMA, Asif Ali KHAN, Rao Sohail Ahmad KHAN. 2022. Expression profiling of transgenes (Cry1Ac and Cry2A) in cotton genotypes under different genetic backgrounds. Journal of Integrative Agriculture, 21(10): 2818-2832.

Adamczyk J J, Meredith J. 2004. Genetic basis for viability of Cry1Ac expression among commercial transgenic Bacillus thuringiensis (Bt) cotton cultivars in the Unites States. Journal of Insect Science, 8, 17–23.
Adamczyk J J, Sumerford D V. 2001. Potential factors impacting season-long expression of Cry1Ac in 13 commercial varieties of Bollgard cotton. Journal of Insect Science, 1, 1–6.
Ahmad S, Cheema H M N, Khan A A, Khan R S A, Ahmad J N. 2019. Resistance status of Helicoverpaarmigera against Bt cotton in Pakistan. Transgenic Research, 28, 199–212.
Ahmad T A, Malik T A, khan I A, Jaskani M J. 2009. Genetic analysis of some morpho-physiological traits related to drought stress. International Journal of Agricultural of Biology, 11, 235–240.
Akhtar Z R, Anjum A A, Saeed Z, Khalid J. 2018. Resistance development in bollworms against Bt proteins deployed in genetically modified cotton. Journal of Entomology and Zoology Studies, 6, 1260–1264.
Ali M A, Bhatti M F, Abbas A, Khan I A. 2010. Assessment of inheritance pattern of some multigenic characters in cotton (G. hirsutum L.). Journal of Agricultural Research, 48, 25–33.
Arshad M, Khan H A A, Rehman M A, Saeed N A. 2015. Incidence of insect predators and parasitoids on transgenic Bt cotton in comparison to non-Bt cotton varieties. Pakistan Journal of Zoology, 47, 823–829.
Bakhsh A, Rao A Q, Shahid A A, Husnain T, Riazuddin S. 2010. CaMV 35S is a developmental promoter being temporal and spatial in expression pattern of insecticidal genes (Cry1Ac and Cry2A) in cotton. Australian Journal of Basic and Applied Sciences, 4, 37–44. 
Banerjee R, Hasler J, Meagher R, Nagoshi R, Hietala L, Huang F, Narva K, Jurat-Fuentes J L. 2017. Mechanism and DNA-based detection of field-evolved resistance to transgenic Bt corn in fall armyworm (Spodoptera frugiperda). Scientific Reports, 7, 10877–10879.
Cheema H M N, Khan A A, Aslam U, Iqrar R, Khan I A. 2016. Assessment of Bt cotton genotypes for Cry1Ac transgene and its expression. Journal of Agricultural Science, 154, 109–117.
Downes S, Walsh T, Tay W T. 2016. Bt resistance in Australian insect pest species. Current Opinion in Insect Science, 15, 78–83.
Guo R Q, Sun S C, Liu B A. 2016. Difference in leaf water use efficiency/photosynthetic nitrogen use efficiency of Bt-cotton and its conventional peer. Scientific Reports, 6, doi: 10.1038/srep33539.
Guo W Z, Sun J, Guo Y F, Zhang T Z. 2001. Investigation of different dosage of inserted Bt genes and their insect resistance in transgenic Bt cotton. Acta Genetica Sinica, 28, 668–676. (in Chinese)
Hussain S S. 2012. Genetic transformation of cotton with Galanthus Nivalis Agglutinin (GNA) gene. Ph D thesis, University of Punjab, Lahore.
ISAAA (International Service for the Acquisition of Agri-biotech Applications). 2017. Global status of commercialized biotech/GM crops in 2017: Biotech crop adoption surges as economic benefits accumulate in 22 years. ISAAA Brief No. 54. Ithaca, NY.
Jabbar A, Mallick S. 1994. Pesticide and environmental situation in Pakistan. Sustainable Development Policy Institute (SDPI), Islamabad, Pakistan. Working Paper Series No. 19.
James C. 2004. Global status of commercialized biotech/GM crops: 2004. ISAAA Brief No. 45. Ithaca, NY.
James C. 2014. Global status of commercialized biotech/GM Crops: 2014. ISAAA Brief No. 49. Ithaca, NY.
Jamil S, Arshad S, Kanwal S, Razzaq H, Shahzad R. 2019. Impact of transgenic crops on global food security: A review. Journal of Agricultural Research, 57, 245–248.
Jamil S, Shahzad R, Iqbal M Z, Yasmeen E, Rahman S U. 2021a. DNA fingerprinting and genetic diversity assessment of GM cotton genotypes for protection of plant breeders rights. International Journal of Agriculture and Biology, 52, 768–776.
Jamil S, Shahzad R, Rahman S U, Iqbal M Z, Yaseen M, Ahmad S, Fatima R. 2021b. The level of Cry1Ac endotoxin and its efficacy against H. armigera in Bt cotton at large scale in Pakistan. GM Crops and Food, 12, 1–17.
Khan G. A, Bakhsh A, Riazuddin S, Husnain T. 2011. Introduction of Cry1Ab gene into cotton (G. hirsutum L.) enhances resistance against lepidopteran pest (Helicoverpa armigera). Spanish Journal of Agricultural Research, 9, 296–300.
Khan M I, Khan A A, Cheema H M N, Khan R S A. 2018. Spatio-temporal and intra-plant expression variability of insecticidal gene (Cry1Ac) in upland cotton. International Journal of Agricultural Biology, 20, 715–722.
Kranthi K R, Naidu S, Dhawad C S, Tatwawadi A, Mate K, Patil E, Bharose A A, Behere G T, Wadaskar R M, Kranthi S. 2005. Temporal and intra-plant variability of Cry1Ac expression in Bt-cotton and its influence on the survival of the cotton bollworm, Helicoverpa armigera (Hubner) (Noctuidae: Lepidoptera). Current Science, 89, 291–298.
Malik T H, Ahsan M Z. 2016. Review of the cotton market in Pakistan and its future prospects. OCL-Oil Seeds and Fats, Crops and Lipids, 23, doi: 10.1051/ocl/2016043.
Memon Q U, Wagan S A, Dong C Y, Xiao S X, Luan J D, Damalas C A. 2019. Health problems from pesticide exposure and personal protective measures among women cotton workers in southern Pakistan. Science of the Total Environment, 685, 659–660.
Nagappa B H, Khadi B M. 2018. Expression of Bt gene (Cry1Ac) on different plant parts at different stages in Bt cotton genotypes. International Journal of Current Microbiology of Applied Science, 7, 3339–3353.
NBC (National Biosafety Committee). 1999. Biosafety Guidelines in Genetic Engineering and Biotechnology. Ministry of Environment Local Government, Rural Development, Government of Pakistan.
Pan Z, Xu L, Zhu Y, Shi H, Chen Z, Chen M, Liu B. 2014. Characterization of a new Cry2A gene of Bacillus thuringiensis with high insecticidal activity against Plutella xylostella L. World Journal of Microbiology and Biotechnology, 30, 2655–2662.
Qamar Z, Tariq M, Rehman T. 2019. Trackable Cemb-Klean cotton transgenic technology: Affordable climate neutral agri-biotech industrialization for developing countries, international. Journal of Biological Science, 6, 131–138.
Qayyum M A, Wakil W, Arif M J, Sahi S T. 2015. Bacillus thuringiensis and Nuclear Polyhedrosis Virus for the enhanced Bio-control of Helicoverpa armigera. International Journal of Agriculture and Biology, 17, 1043–1048.
Saeed M F, Shaheen M, Ahmad I, Zakir A, Nadeem M, Chishti A A, Shahid M, Bakhsh K, Damalas C A. 2017. Pesticide exposure in the local community of Vehari District in Pakistan: an assessment of knowledge and residues in human blood. Science of Total Environment, 147, 587–588.
Shahzad R, Jamil S, Ahamd S, Nisar A, Amina Z, Saleem S, Iqbal M Z, Atif R M, Wang X K. 2021. Harnessing the potential of plant transcription factors in developing climate resilient crops to improve global food security: Current and future perspectives. Saudi Journal of Biological Sciences, 28, 2323–2341.
Shaukat A, Mehmood K, Shaukat I, Naeem M A, Mehfooz A, Saleem M A, Sindhu Z U D, Rajput S A, Hassan M, Umar S, Jamil M A. 2019. Prevalence, haematological alterations and chemotherapy of bovine anaplasmosis in Sahiwal and crossbred cattle of district Faisalabad, Punjab, Pakistan. Pakistan Journal of Zoology, 51, 142–147.
Shera S P, Arora R. 2016. Survival and development of spotted bollworm, Eariasvittella (Fabricius) (Lepidoptera: Nolidae) on different transgenic Bt and isogenic non-Bt cotton genotypes. Phytoparasitica, 44, 99–113.
Spielman D J, Zaidi F, Zambrano P, Khan P A A, Ali S, Cheema H M N. 2017. What are farmers really planting? Measuring the presence and effectiveness of Bt cotton in Pakistan.  PLoS ONE, 12, doi: 10.1371/journal.pone.0176592.
Tabashnik B E, Carriere Y. 2017. Surge in insect resistance to transgenic crops and prospects for sustainability. Journal of Nature Biotechnology, 35, 926–935.
Wan P, Zhang Y, Wu K, Huang M. 2005. Seasonal expression profiles of insecticidal protein and control efficacy against Helicoverpa armigera for Bt cotton in the Yangtze River valley of China. Journal of Economic Entomology, 98, 195–201.
Wu K, Guo Y, Greenplate J T, Deaton R. 2003. Efficacy of transgenic cotton containing Cry1Ac gene from Bacillus thuringiensis L. against Helicoverpa armigera in northern China. Journal of Economic Entomology, 96, 1322–1328.

[1] JIN Yong-mei, MA Rui, YU Zhi-jing, LIN Xiu-feng. Transgenic japonica rice expressing the cry1C gene is resistant to striped stem borers in Northeast China[J]. >Journal of Integrative Agriculture, 2021, 20(11): 2837-2848.
No Suggested Reading articles found!