Pinellia ternata agglutinin expression in chloroplasts confers broad spectrum resistance against aphid, whitefly, Lepidopteran insects, bacterial and viral pathogens

Broad spectrum protection against different insects and pathogens requires multigene engineering. However, such broad spectrum protection against biotic stress is provided by a single protein in some medicinal plants. Therefore, tobacco chloroplasts were transformed with the agglutinin gene from Pinellia ternata (pta), a widely cultivated Chinese medicinal herb. Pinellia ternata agglutinin (PTA) was expressed up to 9.2% of total soluble protein in mature leaves. Purified PTA showed similar hemagglutination activity as snowdrop lectin. Artificial diet with purified PTA from transplastomic plants showed marked and broad insecticidal activity. In planta bioassays conducted with T0 or T1 generation PTA lines showed that the growth of aphid Myzus persicae (Sulzer) was reduced by 89%92% when compared with untransformed (UT) plants. Similarly, the larval survival and total population of whitefly (Bemisia tabaci) on transplastomic lines were reduced by 91%93% when compared with UT plants. This is indeed the first report of lectin controlling whitefly infestation. When transplastomic PTA leaves were fed to corn earworm (Helicoverpa zea), tobacco budworm (Heliothis virescens) or the beet armyworm (spodoptera exigua), 100% mortality was observed against all these three insects. In planta bioassays revealed Erwinia population to be 10 000-fold higher in control than in PTA lines. Similar results were observed with tobacco mosaic virus (TMV) challenge. Therefore, broad spectrum resistance to homopteran (sap-sucking), Lepidopteran insects as well as anti-bacterial or anti-viral activity observed in PTA lines provides a new option to engineer protection against biotic stress by hyper-expression of an unique protein that is naturally present in a medicinal plant

In silico comparative analysis of EST-SSRs in three cotton genomes

In this study, expressed sequence tags- simple sequence repeat (EST-SSRs) were surveyed in three cotton genomes (Gossypium arboreum, Ga; Gossypium raimondii, Gr and Gossypium hirsutum, Gh). The frequency of EST-SSRs was highest in Gr, and motif type for hexanucleotide was obviously abundant in Gr. Trinucleotide repeats were the most abundant motif; AT and AG, AAG and ATC were the most frequent motifs for dinucleotide and trinucleotide, respectively. The repeat number was greatly diverse between the three genomes with the highest variation in Gh. AG and AAG had a high frequency both in homologue groups (HGs) with and without repeat number change between genomes. The range of repeat number change in each HG was wider in Gr-Gh. The annotation of the SSR-ESTs showed that more Gene Ontology (GO) items targeted by SSR-ESTs of Ga and Gr than those of Gh. This study gave us new insights into the difference between the three cotton genomes, which will be more helpful to understand the differentiation and evolution of the three genomes.Key words: Cotton, simple sequence repeat, expressed sequence tags, motif, gene ontology

Engineered Chloroplast dsRNA Silences Cytochrome p450 Monooxygenase, V‐ATPase and Chitin Synthase Genes in the Insect Gut and Disrupts Helicoverpa Armigera Larval Development and Pupation

In the past two decades, chloroplast genetic engineering has been advanced to achieve high‐level protein accumulation but not for down‐regulation of targeted genes. Therefore, in this report, lepidopteran chitin synthase (Chi), cytochrome P450 monooxygenase (P450) and V‐ATPase dsRNAs were expressed via the chloroplast genome to study RNA interference (RNAi) of target genes in intended hosts. PCR and Southern blot analysis confirmed homoplasmy and site‐specific integration of transgene cassettes into the chloroplast genomes. Northern blots and real‐time qRT‐PCR confirmed abundant processed and unprocessed dsRNA transcripts (up to 3.45 million copies of P450 dsRNAs/μg total RNA); the abundance of cleaved dsRNA was greater than the endogenous psbA transcript. Feeding of leaves expressing P450,Chi and V‐ATPase dsRNA decreased transcription of the targeted gene to almost undetectable levels in the insect midgut, likely after further processing of dsRNA in their gut. Consequently, the net weight of larvae, growth and pupation rates were significantly reduced by chloroplast‐derived dsRNAs. Taken together, successful expression of dsRNAs via the chloroplast genome for the first time opens the door to study RNA interference/processing within plastids. Most importantly, dsRNA expressed in chloroplasts can be utilized for gene inactivation to confer desired agronomic traits or for various biomedical applications, including down‐regulation of dysfunctional genes in cancer or autoimmune disorders, after oral delivery of dsRNA bioencapsulated within plant cells

Route to stabilize cubic gauche polynitrogen to ambient conditions via surface-saturation by hydrogen

Cubic gauche polynitrogen (cg-N) is an attractive high-energy density material. However, high-pressure synthesized cg-N will decompose at low-pressure and cannot exist at ambient conditions. Here, the stabilities of cg-N surfaces with and without saturations at different pressures and temperatures are investigated systematically. Pristine surfaces at 0 GPa are very brittle and will decompose at 300 K, especially (1 1 0) surface will collapse completely just after structural relaxation, whereas the decompositions of surfaces can be suppressed by applying pressure, indicating that surface instability causes the cg-N decomposition at low-pressure. Due to the saturation of dangling bonds and transferring electrons to the surfaces, saturation with H can stabilize surfaces at ambient conditions, while OH saturation cannot because of getting electrons from the surfaces. An acidic environment or surface saturation with less electronegative adsorbates is more favorable for the stability of polymerized nitrogen.Comment: 21 pages, 5 figure

Transcriptome Analysis Reveals a Comprehensive Insect Resistance Response Mechanism in Cotton to Infestation by the Phloem Feeding Insect Bemisia Tabaci (Whitefly)

The whitefly (Bemisia tabaci) causes tremendous damage to cotton production worldwide. However, very limited information is available about how plants perceive and defend themselves from this destructive pest. In this study, the transcriptomic differences between two cotton cultivars that exhibit either strong resistance (HR) or sensitivity (ZS) to whitefly were compared at different time points (0, 12, 24 and 48 h after infection) using RNA‐Seq. Approximately one billion paired‐end reads were obtained by Illumina sequencing technology. Gene ontology and KEGG pathway analysis indicated that the cotton transcriptional response to whitefly infestation involves genes encoding protein kinases, transcription factors, metabolite synthesis, and phytohormone signalling. Furthermore, a weighted gene co‐expression network constructed from RNA‐Seq datasets showed that WRKY40 and copper transport protein are hub genes that may regulate cotton defenses to whitefly infestation. Silencing GhMPK3 by virus‐induced gene silencing (VIGS) resulted in suppression of the MPK‐WRKY‐JA and ET pathways and lead to enhanced whitefly susceptibility, suggesting that the candidate insect resistant genes identified in this RNA‐Seq analysis are credible and offer significant utility. Taken together, this study provides comprehensive insights into the cotton defense system to whitefly infestation and has identified several candidate genes for control of phloem‐feeding pests

Identification of miRNAs and their targets by high-throughput sequencing and degradome analysis in cytoplasmic male-sterile line NJCMS1A and its maintainer NJCMS1B of soybean

Table S1. Summary of small RNA annotations from NJCMS1A and NJCMS1B. Table S2. Known miRNAs identified in NJCMS1A and NJCMS1B. Table S3. Family member distribution in conserved miRNA families. Table S4. Summary of miRNA families found in NJCMS1A and NJCMS1B. Table S5. Novel miRNAs on the other arm of known pre-miRNAs. Table S6. Novel miRNAs identified in NJCMS1A and NJCMS1B. Table S7-1. High-confidence known miRNAs identified in NJCMS1A and NJCMS1B. Table S7-2. High-confidence novel miRNAs identified in NJCMS1A and NJCMS1B. Table S8-1. The up-regulated miRNAs identified in NJCMS1A and NJCMS1B. Table S8-2. The down-regulated miRNAs identified in NJCMS1A and NJCMS1B. Table S9. The targets of miRNAs identified in NJCMS1A and NJCMS1B. Table S10. Targets of novel miRNAs in NJCMS1A and NJCMS1B. Table S11. Primers used in this study. (ZIP 637 kb