Jiarui Li, Ph.D., MBA

The soybean cyst nematode (SCN), Heterodera glycines, is one of the most important pests limiting soybean production worldwide. Novel approaches to managing this pest have focused on gene silencing of target nematode sequences using RNA interference (RNAi). With the discovery of endogenous microRNAs as a mode of gene regulation in plants, artificial microRNA (amiRNA) methods have become an alternative method for gene silencing, with the advantage that they can lead to more specific silencing of… Show more

The soybean cyst nematode (SCN), Heterodera glycines, is one of the most important pests limiting soybean production worldwide. Novel approaches to managing this pest have focused on gene silencing of target nematode sequences using RNA interference (RNAi). With the discovery of endogenous microRNAs as a mode of gene regulation in plants, artificial microRNA (amiRNA) methods have become an alternative method for gene silencing, with the advantage that they can lead to more specific silencing of target genes than traditional RNAi vectors. To explore the application of amiRNAs for improving soybean resistance to SCN, three nematode genes (designated as J15, J20, and J23) were targeted using amiRNA vectors. The transgenic soybean hairy roots, transformed independently with these three amiRNA vectors, showed significant reductions in SCN population densities in bioassays. Expression of the targeted genes within SCN eggs were downregulated in populations feeding on transgenic hairy roots. Our results provide evidence that host-derived amiRNA methods have great potential to improve soybean resistance to SCN. This approach should also limit undesirable phenotypes associated with off-target effects, which is an important consideration for commercialization of transgenic crops. Show less

Gall midges induce formation of host nutritive cells and alter plant metabolism to utilize host resources. Here we show that the gene Mayetiola destructor susceptibility-1 on wheat chromosome 3AS encodes a small heat-shock protein and is a major susceptibility gene for infestation of wheat by the gall midge M. destructor, commonly known as the Hessian fly. Transcription of Mayetiola destructor susceptibility-1 and its homoeologs increases upon insect infestation. Ectopic expression of Mayetiola… Show more

Gall midges induce formation of host nutritive cells and alter plant metabolism to utilize host resources. Here we show that the gene Mayetiola destructor susceptibility-1 on wheat chromosome 3AS encodes a small heat-shock protein and is a major susceptibility gene for infestation of wheat by the gall midge M. destructor, commonly known as the Hessian fly. Transcription of Mayetiola destructor susceptibility-1 and its homoeologs increases upon insect infestation. Ectopic expression of Mayetiola destructor susceptibility-1 or induction by heat shock suppresses resistance of wheat mediated by the resistance gene H13 to Hessian fly. Silencing of Mayetiola destructor susceptibility-1 by RNA interference confers immunity to all Hessian fly biotypes on normally susceptible wheat genotypes. Mayetiola destructor susceptibility-1-silenced plants also show reduced lesion formation due to infection by the powdery mildew fungus Blumeria graminis f. sp. tritici. Modification of susceptibility genes may provide broad and durable sources of resistance to Hessian fly, B. graminis f. sp. tritici, and other pests. Show less

Plant-parasitic nematodes are primary biotic factors limiting the crop production. Current nematode control strategies include nematicides, crop rotation and resistant cultivars, but each has serious limitations. RNA interference (RNAi) represents a major breakthrough in the application of functional genomics for plant-parasitic nematode control. RNAi-induced suppression of numerous genes essential for nematode development, reproduction or parasitism has been demonstrated, highlighting the… Show more

Plant-parasitic nematodes are primary biotic factors limiting the crop production. Current nematode control strategies include nematicides, crop rotation and resistant cultivars, but each has serious limitations. RNA interference (RNAi) represents a major breakthrough in the application of functional genomics for plant-parasitic nematode control. RNAi-induced suppression of numerous genes essential for nematode development, reproduction or parasitism has been demonstrated, highlighting the considerable potential for using this strategy to control damaging pest populations. In an effort to find more suitable and effective gene targets for silencing, researchers are employing functional genomics methodologies, including genome sequencing and transcriptome profiling. Microarrays have been used for studying the interactions between nematodes and plant roots and to measure both plants and nematodes transcripts. Furthermore, laser capture microdissection has been applied for the precise dissection of nematode feeding sites (syncytia) to allow the study of gene expression specifically in syncytia. In the near future, small RNA sequencing techniques will provide more direct information for elucidating small RNA regulatory mechanisms in plants and specific gene silencing using artificial microRNAs should further improve the potential of targeted gene silencing as a strategy for nematode management. Show less

To control Heterodera glycines Ichinohe (soybean cyst nematode) in Glycine max (L.) Merr. (soybean), we evaluated the use of producing transgenic soybean seedlings expressing small interfering RNAs (siRNAs) against specific H. glycines genes. Gene fragments of three genes related to nematode reproduction or fitness (Cpn-1, Y25 and Prp-17) were PCR-amplified using specific primers and independently cloned into the pANDA35HK RNAi vector using a Gateway cloning strategy. Soybean roots were… Show more

To control Heterodera glycines Ichinohe (soybean cyst nematode) in Glycine max (L.) Merr. (soybean), we evaluated the use of producing transgenic soybean seedlings expressing small interfering RNAs (siRNAs) against specific H. glycines genes. Gene fragments of three genes related to nematode reproduction or fitness (Cpn-1, Y25 and Prp-17) were PCR-amplified using specific primers and independently cloned into the pANDA35HK RNAi vector using a Gateway cloning strategy. Soybean roots were transformed with these constructions using a composite plant system. Confirmation of transformation was attained by PCR and Southern blot analysis. Transgene expression was detected using reverse transcription PCR (RT-PCR) and expression of siRNAs was confirmed in transgenic plants using northern blot analysis. Bioassays performed on transgenic composite plants expressing double-stranded RNA fragments of Cpn-1, Y25 and Prp-17 genes resulted in a 95, 81 and 79% reduction for eggs g−1 root, respectively. Furthermore, we demonstrated a significant reduction in transcript levels of the Y25 and Prp-17 genes of the nematodes feeding on the transgenic roots via real-time RT-PCR whereas the expression of non-target genes were not affected. The results of this study demonstrate that over-expression of RNA interference constructs of nematode reproduction or fitness-related genes can effectively control H. glycines infection with levels of suppression comparable to conventional resistance. Show less

Production of stable transgenic plants is one factor that limits rapid evaluation of tissue specific transgene expression. To hasten the assessment of transgenes in planta, we evaluated the use of chimeric soybean seedlings expressing transgenic products in roots. Tap roots from four-day old seedlings (cultivars 'Jack' and KS4704) were excised and hairy roots were induced from hypocotyls via Agrobacterium rhizogenes-mediated transformation. Inoculated hypocotyls were screened on a MS-based… Show more

Production of stable transgenic plants is one factor that limits rapid evaluation of tissue specific transgene expression. To hasten the assessment of transgenes in planta, we evaluated the use of chimeric soybean seedlings expressing transgenic products in roots. Tap roots from four-day old seedlings (cultivars 'Jack' and KS4704) were excised and hairy roots were induced from hypocotyls via Agrobacterium rhizogenes-mediated transformation. Inoculated hypocotyls were screened on a MS-based medium containing either 200 mg/L kanamycin or 20 mg/L hygromycin. Beta-glucuronidase (GUS) activity assay indicated that highest GUS expression was observed in hypocotyls exposed to a 4-d pre-inoculation time, a neutral pH (7.0) for the co-cultivation medium. A 170-bp of the Fib-1 gene and 292-bp of the Y25C1A.5 gene fragments, both related to nematode reproduction and fitness, were cloned independently into pANDA35HK vector using a Gateway cloning strategy. The resulting RNAi constructs of the genes fragments were transformed into soybean using the chimeric hairy root system and evaluated for its effect on soybean cyst nematode (Heterodera glycines) fecundity. Confirmation of transformation was attained by polymerase chain reaction and Southern-blot analysis, and some potential for suppression of H. glycines reproduction was detected for the two constructs. This method takes on average four weeks to produce chimeric plants ready for transgene analys Show less