Heterozygosity analysis of spontaneous 2n female gametes and centromere mapping of the diploid Hevea brasiliensis based on full-sib triploid populations.

KEY MESSAGE: Unreduced megagametophytes via second-division restitution were confirmed through heterozygosity analysis, and four candidate physical centromeres of rubber were located for the first time. The evaluation of maternal heterozygosity restitution (MHR) is vital in identifying the mechanism of 2n gametogenesis and assessing the utilization value of 2n gametes. In this study, three full-sib triploid populations were employed to evaluate the MHR of 2n female gametes of rubber tree clone GT1 and to confirm their genetic derivation. The 2n female gametes of GT1 were derived from second-division restitution (SDR) and transmitted more than half of the parental heterozygosity. In addition, low recombination frequency markers were developed, and four candidate physical centromeres of rubber tree were located for the first time. The confirmation that 2n female gametes of rubber tree clone GT1 are derived from SDR provides insights into the molecular mechanisms of 2n gametogenesis. In addition, the identified centromere location will aid in the development of centromeric markers for the rapid identification of the 2n gametogenesis mechanism.

Flower-inducing technology facilitates speed breeding in cassava.

Cassava is a tropical crop that provides daily carbohydrates to more than 800 million people. New cassava cultivars with improved yield, disease resistance, and food quality are critical to end hunger and reduce poverty in the tropics. However, the progress of new cultivar development has been dragged down by difficulties obtaining flowers from desired parental plants to enable designed crosses. Inducing early flowering and increasing seed production are crucial to improving the efficiency of developing farmer-preferred cultivars. In the present study, we used breeding progenitors to evaluate the effectiveness of flower-inducing technology, including photoperiod extension, pruning, and plant growth regulators. Photoperiod extension significantly reduced the time to flowering in all 150 breeding progenitors, especially late-flowering progenitors which were reduced from 6-7 months to 3-4 months. Seed production was increased by using the combination of pruning and plant growth regulators. Combining photoperiod extension with pruning and the PGR 6-benzyladenine (synthetic cytokinin) produced significantly more fruits and seeds than only photoperiod extension and pruning. Another growth regulator, silver thiosulfate, commonly used to block the action of ethylene, did not show a significant effect on fruit or seed production when combined with pruning. The present study validated a protocol for flower induction in cassava breeding programs and discussed factors to consider in implementing the technology. By inducing early flowering and increasing seed production, the protocol helped move one step further for speed breeding in cassava.

Genome-Wide Identification and Analysis of Small Nucleolar RNAs and Their Roles in Regulating Latex Regeneration in the Rubber Tree (Hevea brasiliensis).

Small nucleolar RNAs (snoRNAs) are a class of conserved nuclear RNAs that play important roles in the modification of ribosomal RNAs (rRNAs) in plants. In rubber trees, rRNAs are run off with latex flow during tapping and need to be regenerated for maintaining the functions of the laticifer cells. SnoRNAs are expected to play essential roles in the regeneration of rRNAs. However, snoRNAs in the rubber tree have not been sufficiently characterized thus far. In this study, we performed nuclear RNA sequencing (RNA-seq) to identify snoRNAs globally and investigate their roles in latex regeneration. We identified a total of 3,626 snoRNAs by computational prediction with nuclear RNA-seq data. Among these snoRNAs, 50 were highly expressed in latex; furthermore, the results of reverse transcription polymerase chain reaction (RT-PCR) showed the abundant expression of 31 of these snoRNAs in latex. The correlation between snoRNA expression and adjusted total solid content (TSC/C) identified 13 positively yield-correlated snoRNAs. To improve the understanding of latex regeneration in rubber trees, we developed a novel insulated tapping system (ITS), which only measures the latex regenerated in specific laticifers. Using this system, a laticifer-abundant snoRNA, HbsnoR28, was found to be highly correlated with latex regeneration. To the best of our knowledge, this is the first report to globally identify snoRNAs that might be involved in latex regeneration regulation and provide new clues for unraveling the mechanisms underlying the regulation of latex regeneration.

Production of egg yolk antibody (IgY) against recombinant porcine epidemic diarrhea virus COE protein

Background: Porcine epidemic diarrhea (PED) is a highly contagious disease of pigs, and is characterized by a series ofclinical symptoms, such as severe diarrhea, vomiting and dehydration. Partial protective antigen gene (COE gene) of Sprotein possessing the main B cell epitope, is able to encode proteins with reactogenicity to induce the production of neutralizing antibodies. IgY was found to reduce the mortality in piglets after challenge exposures. Anti-COE IgY antibodyhas never been reported before, here it is described a method for the production of anti-COE IgY, which could be appliedin the treatment for the porcine epidemic diarrhea virus (PEDV) infection.Materials, Methods & Results: A PEDV strain was isolated from a clinical sample. The COE ORF (Open reading frame,ORF)was amplifi ed by PCR and iserted into the pMD18-T clone vector. The isolated was defi ned as Porcine epidemic diarrheavirus strain JS-HZ2012 subtype by sequencing, the clinical sample was defi ned as the nucleic acid sequence has a 99.5%homology with that of PEDV CV777 strain. And then the COE ORF was subcloned into pET-32a by T4 DNA ligase andintroduced into the E.coli Bal21 (DE3). COE protein was produced by the induction of the E.coli Bal21 containing pET32a-COE with isopropyl-β-D-1-thiogalactopyrannoside (IPTG). Expression of the recombinant COE protein (rCOE)fused with His-tag was analyzed by SDS-PAGE and detected by western-blotting using anti-His monoclonal antibody.The rCOE was purifi ed by Ni+ affi nity purifi cation chromatography under denature condition and dialyzed against PBS.The concentration of the rCOE was determined by BCA method. After immunnizing the chickens with rCOE , All animalhandling procedures were performed under veterinary supervision and following the recommendations of the local lawsand regulations on Animal Experimentation. Anti-COE IgY was isolated by chloroform extraction and...

Production of egg yolk antibody (IgY) against recombinant porcine epidemic diarrhea virus COE protein

Background: Porcine epidemic diarrhea (PED) is a highly contagious disease of pigs, and is characterized by a series ofclinical symptoms, such as severe diarrhea, vomiting and dehydration. Partial protective antigen gene (COE gene) of Sprotein possessing the main B cell epitope, is able to encode proteins with reactogenicity to induce the production of neutralizing antibodies. IgY was found to reduce the mortality in piglets after challenge exposures. Anti-COE IgY antibodyhas never been reported before, here it is described a method for the production of anti-COE IgY, which could be appliedin the treatment for the porcine epidemic diarrhea virus (PEDV) infection.Materials, Methods & Results: A PEDV strain was isolated from a clinical sample. The COE ORF (Open reading frame,ORF)was amplifi ed by PCR and iserted into the pMD18-T clone vector. The isolated was defi ned as Porcine epidemic diarrheavirus strain JS-HZ2012 subtype by sequencing, the clinical sample was defi ned as the nucleic acid sequence has a 99.5%homology with that of PEDV CV777 strain. And then the COE ORF was subcloned into pET-32a by T4 DNA ligase andintroduced into the E.coli Bal21 (DE3). COE protein was produced by the induction of the E.coli Bal21 containing pET32a-COE with isopropyl-β-D-1-thiogalactopyrannoside (IPTG). Expression of the recombinant COE protein (rCOE)fused with His-tag was analyzed by SDS-PAGE and detected by western-blotting using anti-His monoclonal antibody.The rCOE was purifi ed by Ni+ affi nity purifi cation chromatography under denature condition and dialyzed against PBS.The concentration of the rCOE was determined by BCA method. After immunnizing the chickens with rCOE , All animalhandling procedures were performed under veterinary supervision and following the recommendations of the local lawsand regulations on Animal Experimentation. Anti-COE IgY was isolated by chloroform extraction and...(AU)