Plant-mediated RNAi has been developed as a powerful weapon in the fight against agricultural insect pests. The gap gene hunchback (hb) is of crucial importance in insect axial patterning and knockdown of hb is deforming and lethal to the next generation. The peach potato aphid, Myzus persicae (Sulzer), has many host plants and can be found throughout the world. To investigate the effect of plant-mediated RNAi on control of this insect, the hb gene in M. persicae was cloned, plant RNAi vector was constructed, and transgenic tobacco expressing Mphb dsRNA was developed. Transgenic tobacco had a different integration pattern of the transgene. Bioassays were performed by applying neonate aphids to homozygous transgenic plants in the T2 generation. Results revealed that continuous feeding of transgenic diet reduced Mphb mRNA level in the fed aphids and inhibited insect reproduction, indicating successful knockdown of the target gene in M. persicae by plant-mediated RNAi.

The gap gene hunchback (hb) is a key regulator in the anteroposterior patterning of insects. Loss-of-function of hb resulted in segmentation defects in the next generation. In this paper, hb expression level was investigated at different developmental stages of the pea aphid, Acyrthosiphon pisum (Ap). Aphb mRNA was most early detected at the first instar stage and showed an incontinuous increase in the whole life cycle. Ingested RNA interference was performed at the second instar stage to knockdown the Aphb expression. Continuous feeding of Aphb double-stranded RNA mixed in artificial diet led to reduction of Aphb transcripts and rise of insect lethality. These results indicated that hunchback was a good RNAi target in the management of insect pests.

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Vitellogenin (Vg) is a precursor of major egg storage protein, vitellin (Vt), and plays primary roles in reproduction of oviparous vertebrates and invertebrates. Chrysopa septempunctata Wesmael is an important and common predator of various insect pests. Here, we first cloned C. septempunctata Vg gene, CsVg. The complete CsVg cDNA was 5664 bp, which encodes an 1810-residues protein with a predicted molecular mass of 206.23 kDa. Expression profile revealed that CsVg mRNA first appeared on day 4 after emergence, maximally accumulated on day 10, and then declined gradually. RNAi mediated by injection of dsRNA depleted CsVg transcripts, significantly reduced egg-laying amount, and decreased egg hatching rate, suggesting that CsVg functions through effects on egg production and hatching in C. septempunctata.

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PebC1, a novel protein elicitor was isolated and purified from the mycelium of gray mold fungus, Botrytis cinerea strain BC-4-2-2-1. The protein was eluted through HiTrap DEAE FF and RESOURCE Q anion exchange chromatography and displayed as a single band with an apparent molecular weight of 36 kDa on silver staining sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The pI of the purified protein PebC1 was determined by 2-DE and was 4.85. Three peptide segments were obtained by MALDI-TOF. Similarity, the homology matching using protein BLAST search found that two proteins, viz. XP_001593856 and XP_001551609 were having high score and covered sequence of the three peptides. Protein XP_001551609, a deduced protein nascent polypeptide-associated complex alpha-polypeptide, was more authentic because it was from Botryotinia fuckeliana that is better known as its anamorph, B.cinerea and showed 95% homology with the three polypeptides. The full cDNA sequence encoding for pebC1 (Genbank accession number FJ748868) was amplified from B. cinerea and consists of 639bp, which is same as a registered gene of XM_001551559, a nascent polypeptide-associated complex alpha-polypeptide partial mRNA. The gene encode a hypothetical protein speculated from an annotated genomic sequence from B. fuckeliana B05.10 (NW_001814507) and there is no publication about the gene. The PebC1 protein significantly promoted wheat seedling growth with an optimum protein concentration of 5 microg/mL. Root systemic activity of wheat with 4-5 leaves increased by 1.29 fold, and the wheat seedling drought resistance integrated index increased from 36.53 to 57.08 under two cycles of drought stress after treatment of PebC1. PebC1 protein at the optimum concentration of 10 microg/mL induced 69.19% disease resistance against gray mold fungus in tomato. Furthermore, phenylalanine ammonia-lyase (PAL), peroxides (POD), and polyphenol oxidase (PPO) related to plant resistance metabolism were also increased considerably after PebC1 treatment. PAL activity was increased by 46.84% at 24h post-treatment, while POD and PPO activity increased by 109.5% and 111.0% at 72 h, respectively over the control.

Systemic acquired resistance (SAR) is an inducible defense mechanism which plays a central role in protecting plants from pathogen attack. A new elicitor, PeaT1 from Alternaria tenuissima, was expressed in Escherichia coil and characterized with systemic acquired resistance to tobacco mosaic virus (TMV). PeaT1-treated plants exhibited enhanced systemic resistance with a significant reduction in number and size of TMV lesions on wild tobacco leaves as compared with control. The quantitative analysis of TMV CP gene expression with real-time quantitative PCR showed there was reduction in TMV virus concentration after PeaT1 treatment. Similarly, peroxidase (POD) activity and lignin increased significantly after PeaT1 treatment. The real-time quantitative PCR revealed that PeaT1 also induced the systemic accumulation of pathogenesis-related gene, PR-1a and PR-1b which are the markers of systemic acquired resistance (SAR), NPR1 gene for salicylic acid (SA) signal transduction pathway and PAL gene for SA synthesis. The accumulation of SA and the failure in development of similar level of resistance as in wild type tobacco plants in PeaT1 treated nahG transgenic tobacco plants indicated that PeaT1-induced resistance depended on SA accumulation. The present work suggested that the molecular mechanism of PeaT1 inducing disease resistance in tobacco was likely through the systemic acquired resistance pathway mediated by salicylic acid and the NPR1 gene.

Aphid, a short germband insect, displays an embryogenesis different from that of long germband insect species. Furthermore, the development of its parthenogenetic and viviparous embryo is different from that of the embryo resulting from sexual reproduction. To better understand the genetic regulation of this type of embryogenesis, the functions of hunchback in asexual Acyrthosiphon pisum were investigated by parental RNAi. Microinjection of Aphb double-stranded RNA yielded several defective phenotypes. Quantitative real-time PCR analysis revealed that these defects resulted from reduction of Aphb mRNA level in injected aphids. All these results suggested that the hb gene in parthenogenetic and viviparous Acyrthosiphon pisum was involved in abdominal identity suppression and germband growth as its homologue does in sexual insects.