Dr. Peng Zhou’s research team in Tropical Bioscience and Biotechnology Research Institute (TBBRI) has made important progress in the technology of molecular vector construction. A new method for rapid construction of plant hpRNA expression vector has been developed by Zhou’s team , which can be used to construct hpRNA expression vectors targeting a variety of plant genes (single or multiple). This method provides a novel and high-throughput platform for large-scale analysis of plant functional genomics.
After double-stranded RNA was discovered as the trigger of RNA interference (RNAi), RNAi has become one of the most powerful tools for the analysis of gene function. Hairpin RNA (hpRNA) constructs are commonly used to induce degradation of target genes through RNAi mechanisms. In plants, intron-containing hairpin RNA (ihpRNA) with an intron as a spacer sequence shows the highest gene silencing efficiency. Therefore, ihpRNA constructs have been widely used for gene silencing in plants. With the explosive release of gene sequences and genomic sequences in plants, a high-throughput and cost-efficient system for making ihpRNA constructs is in great demand. To facilitate the generation of ihpRNA constructs, Zhou’s research team has developed a one-step and cost-effective method via their new plant RNAi vector pRNAi-GG based on Golden Gate cloning.
Using this system, a plant ihpRNA expression vector can be made from a single PCR product of the gene of interest by one-tube restriction-ligation reaction and one-step transformation. This method has been successfully applied to generate ihpRNA constructs for the effective silencing of various individual endogenous and exogenous marker genes as well as two genes simultaneously. Their results suggested that this novel method provides a high-throughput and reliable platform for making ihpRNA constructs.
After publishing, this method has been used by many labs, including more than 30 labs in China, and more than 60 foreign labs so far.
After double-stranded RNA was discovered as the trigger of RNA interference (RNAi), RNAi has become one of the most powerful tools for the analysis of gene function. Hairpin RNA (hpRNA) constructs are commonly used to induce degradation of target genes through RNAi mechanisms. In plants, intron-containing hairpin RNA (ihpRNA) with an intron as a spacer sequence shows the highest gene silencing efficiency. Therefore, ihpRNA constructs have been widely used for gene silencing in plants. With the explosive release of gene sequences and genomic sequences in plants, a high-throughput and cost-efficient system for making ihpRNA constructs is in great demand. To facilitate the generation of ihpRNA constructs, Zhou’s research team has developed a one-step and cost-effective method via their new plant RNAi vector pRNAi-GG based on Golden Gate cloning.
Using this system, a plant ihpRNA expression vector can be made from a single PCR product of the gene of interest by one-tube restriction-ligation reaction and one-step transformation. This method has been successfully applied to generate ihpRNA constructs for the effective silencing of various individual endogenous and exogenous marker genes as well as two genes simultaneously. Their results suggested that this novel method provides a high-throughput and reliable platform for making ihpRNA constructs.
After publishing, this method has been used by many labs, including more than 30 labs in China, and more than 60 foreign labs so far.
