Ectopic expression of the Arabidopsis mutant L3 NB-LRR receptor gene in Nicotiana benthamiana cells leads to cell death

Huang, Jianzhong; Jia, Peng; Zhong, Xiaoju; Guan, Xiuying; Zhang, Hongbin; Gao, Zhiyong

doi:10.1016/j.gene.2024.148256

Cited by 4 publications

(1 citation statement)

References 34 publications

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“…Transient gene expression studies have shown that maximum expression occurs within 18 hours (h) to 48 h after inoculation, and can last for 10 days [5]. Transient expression systems are often used for the analysis of proteins including protein subcellular localization studies, co-immunoprecipitation (Co-IP), and bimolecular fluorescence complementation (BiFC) [6] [7].…”

Section: Introductionmentioning

confidence: 99%

The application ofNicotiana benthamianaas a Transient Expression Host to Clone the Coding Sequences of Plant Genes

Huang

et al. 2022

Preprint

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Coding sequences (CDS) are commonly used for transient gene expression, in yeast two-hybrid screening, to verify protein interaction verification interactions and in prokaryotic gene expression studies. CDS are most common method to obtain CDS is to utilize commonly obtained using complementary DNA (cDNA) as a template, which derived from message RNA (mRNA) extracted from plant tissues and generated by reverse transcription. However, some CDSs are difficult to acquire with through regular way due to the process as they are expressed at extremely low gene expressions levels or have specific spatial and/or temporal expression patterns in vivo, so it is necessary to develop. These challenges require the development of alternative CDS cloning technologies. In this study, we found that the genomic intron-containing gene coding sequences (gDNA) from Arabidopsis thaliana, Oryza sativa, Brassica napus, and Glycine max can be correctly transcribed and spliced into mRNA in Nicotiana benthamiana (N. benthamiana). In contrast, gDNAs from Triticum aestivum and Sorghum bicolor did not function correctly. In transient expression experiments, the target DNA sequence can is driven by a constitutive promoter. Theoretically, a sufficient amount of mRNA can be extracted from the N. benthamiana leaves, which is making it conducive to the cloning of CDS of target genes. Our data demonstrate that N. benthamiana can be employed as an effective host for the cloning CDS of plant genes.

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Section: Introductionmentioning

confidence: 99%

The application ofNicotiana benthamianaas a Transient Expression Host to Clone the Coding Sequences of Plant Genes

Huang

et al. 2022

Preprint

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Genome-wide association study of Verticillium longisporum resistance in Brassica genotypes

Wang,

Fredua-Agyeman,

et al. 2024

Front. Plant Sci.

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Verticillium stripe, caused by Verticillium longisporum, presents an emerging threat to Canadian canola (Brassica napus). Initially detected in Manitoba in 2014, the presence of this pathogen has since been confirmed across western Canada. Infections by V. longisporum can result in yield losses of up to 50%, which is a cause for concern given the susceptibility of most commercial Canadian canola cultivars. The objective of this study was to screen a collection of 211 Brassica genotypes for their reactions to V. longisporum, and to use genome-wide association study (GWAS) to identify single nucleotide polymorphism (SNP) markers for resistance. The plant material consisted of 110 rutabaga (B. napus ssp. napobrassica), 35 canola, 40 Brassica rapa, and 15 Brassica oleracea accessions or cultivars, alongside 11 hosts of the European Clubroot Differential (ECD) set. These materials were screened for resistance under greenhouse conditions and were genotyped using a 19K Brassica SNP array. Three general linear models (GLM), four mixed linear models (MLM), and three GWAS methods were employed to evaluate the markers. Eleven non-commercial Brassica accessions and 9 out of 35 commercial canola cultivars displayed a low normalized area under the disease progress curve (AUDPCnorm.). The non-commercial accessions could prove valuable as potential sources of resistance against V. longisporum. Forty-five SNP markers were identified to be significantly associated with V. longisporum resistance using single-SNP based GWAS analysis. In comparison, haplotype-based GWAS analyses identified 10 to 25 haplotype blocks to be significantly associated with V. longisporum resistance. Between 20% and 56% of QTLs identified by the more conventional single-SNP based GWAS analysis were also detected by the haplotype-based GWAS analysis. The overlapping genomic regions identified by the two GWAS methods present promising hotspots for marker-assisted selection in the future development of Verticillium stripe-resistant canola.

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The Application of <i>Nicotiana benthamiana</i> as a Transient Expression Host to Clone the Coding Sequences of Plant Genes

Huang,

Jia,

Zhong

et al. 2024

AJMB

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Coding sequences (CDS) are commonly used for transient gene expression, in yeast two-hybrid screening, to verify protein interactions and in prokaryotic gene expression studies. CDS are most commonly obtained using complementary DNA (cDNA) derived from messenger RNA (mRNA) extracted from plant tissues and generated by reverse transcription. However, some CDS are difficult to acquire through this process as they are expressed at extremely low levels or have specific spatial and/or temporal expression patterns in vivo. These challenges require the development of alternative CDS cloning technologies. In this study, we found that the genomic intron-containing gene coding sequences (gDNA) from Arabidopsis thaliana, Oryza sativa, Brassica napus, and Glycine max can be correctly transcribed and spliced into mRNA in Nicotiana benthamiana. In contrast, gDNAs from Triticum aestivum and Sorghum bicolor did not function correctly. In transient expression experiments, the target DNA sequence is driven by a constitutive promoter. Theoretically, a sufficient amount of mRNA can be extracted from the N. benthamiana leaves, making it conducive to the cloning of CDS target genes. Our data demonstrate that N. benthamiana can be used as an effective host for the cloning CDS of plant genes.

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Ectopic expression of the Arabidopsis mutant L3 NB-LRR receptor gene in Nicotiana benthamiana cells leads to cell death

Cited by 4 publications

References 34 publications

The application ofNicotiana benthamianaas a Transient Expression Host to Clone the Coding Sequences of Plant Genes

The application ofNicotiana benthamianaas a Transient Expression Host to Clone the Coding Sequences of Plant Genes

Genome-wide association study of Verticillium longisporum resistance in Brassica genotypes

The Application of <i>Nicotiana benthamiana</i> as a Transient Expression Host to Clone the Coding Sequences of Plant Genes

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Ectopic expression of the Arabidopsis mutant L3 NB-LRR receptor gene in Nicotiana benthamiana cells leads to cell death

Cited by 4 publications

References 34 publications

The application ofNicotiana benthamianaas a Transient Expression Host to Clone the Coding Sequences of Plant Genes

The application ofNicotiana benthamianaas a Transient Expression Host to Clone the Coding Sequences of Plant Genes

Genome-wide association study of Verticillium longisporum resistance in Brassica genotypes

The Application of &lt;i&gt;Nicotiana benthamiana&lt;/i&gt; as a Transient Expression Host to Clone the Coding Sequences of Plant Genes

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The Application of <i>Nicotiana benthamiana</i> as a Transient Expression Host to Clone the Coding Sequences of Plant Genes