Near-Infrared Nanophosphor Embedded in Mesoporous Silica Nanoparticle with High Light-Harvesting Efficiency for Dual Photosystem Enhancement.

Light-harvesting and conversion ability is important to promote plant growth, and especially when resources are limited. A near-infrared (NIR) nanophosphor embedded with mesoporous silica nanoparticles (MSN), ZnGa2 O4 :Cr3+ ,Sn4+ (ZGOCS), was developed and its optical properties were harnessed to enhance the photosynthetic ability of Brassica rapa spp. chinensis. The broad excitation of ZGOCS from the ultraviolet to the visible region allowed the conversion of extra light into near-infrared light (650-800 nm) and thus promoted the dual photosystem via the Emerson effect. ZGOCS@MSN was spherical with a size of 65±10 nm and good dispersion. A light conversion ability of up to 75 % under different wavelengths was achieved. Moreover, the electron transfer rate of photosynthesis increased by 100 % with a suitable ZGOCS@MSN concentration. Plant and animal models were used to explore the effects of the nanophosphor. ZGOCS@MSN distribution was tracked by monitoring its NIR emission in plant and animal tissues, demonstrating that this nanophosphor can be potentially utilized in plant growth.

Two Paralogous Genes Encoding Auxin Efflux Carrier Differentially Expressed in Bitter Gourd (Momordica charantia).

The phytohormone auxin regulates various developmental programs in plants, including cell growth, cell division and cell differentiation. The auxin efflux carriers are essential for the auxin transport. To show an involvement of auxin transporters in the coordination of fruit development in bitter gourd, a juicy fruit, we isolated novel cDNAs (referred as McPIN) encoding putative auxin efflux carriers, including McPIN1, McPIN2 (allele of McPIN1) and McPIN3, from developing fruits of bitter gourd. Both McPIN1 and McPIN3 genes possess six exons and five introns. Hydropathy analysis revealed that both polypeptides have two hydrophobic regions with five transmembrane segments and a predominantly hydrophilic core. Phylogenetic analyses revealed that McPIN1 shared the highest homology to the group of Arabidopsis, cucumber and tomato PIN1, while McPIN3 belonged to another group, including Arabidopsis and tomato PIN3 as well as PIN4. This suggests different roles for McPIN1 and McPIN3 in auxin transport involved in the fruit development of bitter gourd. Maximum mRNA levels for both genes were detected in staminate and pistillate flowers. McPIN1 is expressed in a particular period of early fruit development but McPIN3 continues to be expressed until the last stage of fruit ripening. Moreover, these two genes are auxin-inducible and qualified as early auxin-response genes. Their expression patterns suggest that these two auxin transporter genes play a pivotal role in fruit setting and development.

Global Transcriptomic Analysis of Targeted Silencing of Two Paralogous ACC Oxidase Genes in Banana.

Among 18 1-aminocyclopropane-1-carboxylic acid (ACC) oxidase homologous genes existing in the banana genome there are two genes, Mh-ACO1 and Mh-ACO2, that participate in banana fruit ripening. To better understand the physiological functions of Mh-ACO1 and Mh-ACO2, two hairpin-type siRNA expression vectors targeting both the Mh-ACO1 and Mh-ACO2 were constructed and incorporated into the banana genome by Agrobacterium-mediated transformation. The generation of Mh-ACO1 and Mh-ACO2 RNAi transgenic banana plants was confirmed by Southern blot analysis. To gain insights into the functional diversity and complexity between Mh-ACO1 and Mh-ACO2, transcriptome sequencing of banana fruits using the Illumina next-generation sequencer was performed. A total of 32,093,976 reads, assembled into 88,031 unigenes for 123,617 transcripts were obtained. Significantly enriched Gene Oncology (GO) terms and the number of differentially expressed genes (DEGs) with GO annotation were 'catalytic activity' (1327, 56.4%), 'heme binding' (65, 2.76%), 'tetrapyrrole binding' (66, 2.81%), and 'oxidoreductase activity' (287, 12.21%). Real-time RT-PCR was further performed with mRNAs from both peel and pulp of banana fruits in Mh-ACO1 and Mh-ACO2 RNAi transgenic plants. The results showed that expression levels of genes related to ethylene signaling in ripening banana fruits were strongly influenced by the expression of genes associated with ethylene biosynthesis.

Bioconversion of pinoresinol into matairesinol by use of recombinant Escherichia coli.

Lignans, a class of dimeric phenylpropanoid derivative found in plants, such as whole grains and sesame and flax seeds, have anticancer activity and can act as phytoestrogens. The lignans secoisolariciresinol and matairesinol can be converted in the mammalian proximal colon into enterolactone and enterodiol, respectively, which reduce the risk of breast and colon cancer. To establish an efficient bioconversion system to generate matairesinol from pinoresinol, the genes encoding pinoresinol-lariciresinol reductase (PLR) and secoisolariciresinol dehydrogenase (SDH) were cloned from Podophyllum pleianthum Hance, an endangered herb in Taiwan, and the recombinant proteins, rPLR and rSDH, were expressed in Escherichia coli and purified. The two genes, termed plr-PpH and sdh-PpH, were also linked to form two bifunctional fusion genes, plr-sdh and sdh-plr, which were also expressed in E. coli and purified. Bioconversion in vitro at 22°C for 60 min showed that the conversion efficiency of fusion protein PLR-SDH was higher than that of the mixture of rPLR and rSDH. The percent conversion of (+)-pinoresinol to matairesinol was 49.8% using PLR-SDH and only 17.7% using a mixture of rPLR and rSDH. However, conversion of (+)-pinoresinol by fusion protein SDH-PLR stopped at the intermediate product, secoisolariciresinol. In vivo, (+)-pinoresinol was completely converted to matairesinol by living recombinant E. coli expressing PLR-SDH without addition of cofactors.

Oral immunogenicity of porcine reproductive and respiratory syndrome virus antigen expressed in transgenic banana.

Porcine reproductive and respiratory syndrome virus (PRRSV) is a persistent threat of economically significant influence to the swine industry worldwide. Recombinant DNA technology coupled with tissue culture technology is a viable alternative for the inexpensive production of heterologous proteins in planta. Embryogenic cells of banana cv. 'Pei chiao' (AAA) have been transformed with the ORF5 gene of PRRSV envelope glycoprotein (GP5) using Agrobacterium-mediated transformation and have been confirmed. Recombinant GP5 protein levels in the transgenic banana leaves were detected and ranged from 0.021%-0.037% of total soluble protein. Pigs were immunized with recombinant GP5 protein by orally feeding transgenic banana leaves for three consecutive doses at a 2-week interval and challenged with PRRSV at 7 weeks postinitial immunization. A vaccination-dependent gradational increase in the elicitation of serum and saliva anti-PRRSV IgG and IgA was observed. Furthermore, significantly lower viraemia and tissue viral load were recorded when compared with the pigs fed with untransformed banana leaves. The results suggest that transgenic banana leaves expressing recombinant GP5 protein can be an effective strategy for oral delivery of recombinant subunit vaccines in pigs and can open new avenues for the production of vaccines against PRRSV.

A comparison of strategies for multiple-gene co-transformation via hairy root induction.

Hairy root is a transformed root tissue in which transfer DNA (T-DNA) is inserted in the genome by Agrobacterium rhizogenes. To establish a system for multiple-gene co-transformation in hairy roots, we evaluated four different strategies using A. rhizogenes. The genes gusA and mgfp5 were located in separate plasmids, which were transformed into two different batches of A. rhizogenes (strategy 2AR) or a single batch (strategy 2BV). The two reporter genes were also inserted in one T-DNA (strategy 1TD) or two different T-DNAs (strategy 2TD) in a binary vector. Over 90 % of infected Nicotiana tabacum leaf discs formed hairy roots in all four groups, which was not significantly different from the infection efficiency of wild-type A. rhizogenes. Proportions of co-transformed hairy roots with strategies 2AR, 2BV, 1TD, and 2TD were 65.4, 40.0, 78.6, and 82.1 %, respectively, which indicated that all of the strategies were suitable for co-transformation of multiple genes. High variation in growth rate and heterologous protein expression indicated that further screening is required to identify the clone with the highest productivity. Our results indicated that strategies 1TD and 2TD achieved the highest co-transformation efficiency. Combination with strategy 2AR or 2BV provides additional options for co-transformation of multiple transgenes.

Characterization of MicroRNAs and Gene Expression in ACC Oxidase RNA Interference-Based Transgenic Bananas.

Banana (Musa acuminata, AAA group) is a typically respiratory climacteric fruit. Previously, genes encoding ACC oxidase, one of the key enzymes in ethylene biosynthesis, Mh-ACO1 and Mh-ACO2 in bananas were silenced individually using RNAi interference technology, and fruit ripening of transgenic bananas was postponed. Here, the differential expression of miRNAs and their targeted mRNAs were analyzed in the transcriptomes of fruits at the third ripening stage, peel color more green than yellow, from the untransformed and RNAi transgenic bananas. Five significantly differentially expressed miRNAs (mac-miR169a, mac-miR319c-3p, mac-miR171a, mac-miR156e-5p, and mac-miR164a-5p) were identified. The predicted miRNA target genes were mainly enriched in six KEGG pathways, including 'sulfur relay system', 'protein digestion and absorption', 'histidine metabolism', 'pathogenic E. coli infection', 'sulfur metabolism', and 'starch and sucrose metabolism'. After ethylene treatment, the expression of ACC oxidase silencing-associated miRNAs was down-regulated, and that of their target genes was up-regulated along with fruit ripening. The evolutionary clustering relationships of miRNA precursors among 12 gene families related to fruit ripening were analyzed. The corresponding expression patterns of mature bodies were mainly concentrated in flowers, fruits, and leaves. Our results indicated that ethylene biosynthesis is associated with miRNAs regulating the expression of sulfur metabolism-related genes in bananas.

Expression Fluctuations of Genes Involved in Carbohydrate Metabolism Affected by Alterations of Ethylene Biosynthesis Associated with Ripening in Banana Fruit.

The banana is a typical climacteric fruit that undergoes ethylene dependent ripening. During fruit ripening, ethylene production triggers a developmental cascade that results in a series of physiological and biochemical changes. The fruit transcriptomes of untransformated wild-type (WT) and RNAi transgenic banana plants for Mh-ACO1 and Mh-ACO2 have been previously sequenced and analyzed, and most of the differentially expressed genes were enriched in 'carbon fixation in photosynthetic organism', 'cysteine and methionine metabolism', 'citrate cycle (tricarboxylic acid cycle, TCA cycle)', and 'starch and sucrose metabolism' based on Kyoto Encyclopedia of Genes and Genomes (KEGG) annotation. In this research, we investigated the expression fluctuations of genes involved in carbohydrate metabolism affected by alterations of ethylene biosynthesis associated with ripening in banana fruits. Expression profiles of sucrose synthase, sucrose phosphate synthase, neutral invertase, and acidic invertase/ß-fructofuranosidase, as analyzed by Avadis and Trinity, showed that both analyses were complementary and consistent. The overall gene expression tendency was confirmed by the implementation of quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR) with mRNAs of banana fruits in Mh-ACO1 and Mh-ACO2 RNAi transgenic plants. These results indicated that altered expression of genes associated with ethylene biosynthesis strongly influenced the expression levels of genes related to starch and sucrose metabolism, as well as the glycolysis pathway in ripening banana fruits.

Metabolomic assessment of arsenite toxicity and novel biomarker discovery in early development of zebrafish embryos.

CONTEXT: Arsenic poisoning commonly occurs through exposure to water contaminated with arsenic and causes long-term symptoms. Of all the arsenic derivatives, arsenite is the one of the most toxic compounds. However, the toxicity of arsenite during developmental stages is still unclear. OBJECTIVE: In this study, we performed a metabolomic analysis of arsenite responses in embryonic zebrafish. MATERIALS AND METHODS: Embryonic zebrafish were used as an animal model in this study. They were exposed to sodium arsenite under different concentrations (0.5, 1.0, 2.0, and 5.0 mg/L) in 24 h, 48 h and 72 h post fertilization. Changes in morphology were observed through a light microscope. Changes in metabolomics were identified using an ultraperformance liquid chromatography quadrupole time-of-flight system. RESULTS: The IC50 range was 0.75 ±â€¯0.25 mg/L. Compared with the control group, the embryonic lethality rate decreased to 33.3% under 1.0 mg/L of arsenite treatment, whereas it decreased to 20.0% under 2.0 mg/L of arsenite treatment. Numerous body axis curvatures were also observed under treatment with 2.0 and 5.0 mg/L of arsenic. Pericardium and yolk sac edema were randomly discovered and found to worsen over time. Moreover, the 10 metabolites with the highest variable importance in projection score were identified as potential biomarkers for arsenic exposure. CONCLUSION: Arsenic exposure not only leads to a change in the morphology of embryonic zebrafish but also disturbs the metabolism of zebrafish in early developmental stages.

Distinct expression of CDCA3, CDCA5, and CDCA8 leads to shorter relapse free survival in breast cancer patient.

Breast cancer is a dangerous disease that results in high mortality rates for cancer patients. Many methods have been developed for the treatment and prevention of this disease. Determining the expression patterns of certain target genes in specific subtypes of breast cancer is important for developing new therapies for breast cancer. In the present study, we performed a holistic approach to screening the mRNA expression of six members of the cell division cycle-associated gene family (CDCA) with a focus on breast cancer using the Oncomine and The Cancer Cell Line Encyclopedia (CCLE) databases. Furthermore, Gene Expression-Based Outcome for Breast Cancer Online (GOBO) was also used to deeply mine the expression of each CDCA gene in clinical breast cancer tissue and breast cancer cell lines. Finally, the mRNA expression of the CDCA genes as related to breast cancer patient survival were analyzed using a Kaplan-Meier plot. CDCA3, CDCA5, and CDCA8 mRNA expression levels were significantly higher than the control sample in both clinical tumor sample and cancer cell lines. These highly expressed genes in the tumors of breast cancer patients dramatically reduced patient survival. The interaction network of CDCA3, CDCA5, and CDCA8 with their co-expressed genes also revealed that CDCA3 expression was highly correlated with cell cycle related genes such as CCNB2, CDC20, CDKN3, and CCNB1. CDCA5 expression was correlated with BUB1 and TRIP13, while CDCA8 expression was correlated with BUB1 and CCNB1. Altogether, these findings suggested CDCA3, CDCA5, and CDCA8 could have a high potency as targeted breast cancer therapies.

Novel therapeutic effects of sesamin on diabetes-induced cardiac dysfunction.

Diabetes is a risk factor that increases the occurrence and severity of cardiovascular events. Cardiovascular complications are the leading cause of mortality of 75% of patients with diabetes >40 years old. Sesamin, the bioactive compound extracted from Sesamum indicum, is a natural compound that has diverse beneficial effects on hypoglycemia and reducing cholesterol. The aim of this study is to investigate sesamin effects to diabetes-inducing cardiac hypertrophy. In the present study bioinformatics analysis demonstrated cardiac hypertrophy signaling may be the most important pathway for upregulating genes in sesamin-treated groups. To verify the bioinformatics prediction, sesamin was used as the main bioactive compound to attenuate the impact of diabetes induced by streptozotocin (STZ) on cardiac function in a rat model. The results revealed that oral administration of sesamin for 4 weeks (100 and 200 mg/kg body weight) marginally improved blood glucose levels, body weight and significantly ameliorated the effects on heart rate and blood pressure in rats with type 1 diabetes relative to control rats. The QT interval of sesamin was also reduced relative to the control group. The findings indicated that sesamin has potential cardioprotective effects in the STZ-induced diabetes model. This suggested that this can be used as a novel treatment for patients with diabetes with cardiac dysfunction complication.

Genomic organization of a diverse ACC synthase gene family in banana and expression characteristics of the gene member involved in ripening of banana fruits.

The banana is one of the typical climacteric fruits with great economic importance in agriculture. To understand the basic mechanism underlying banana ripening, gene clones for banana ACC synthase (EC 4.4.1.14), a key regulatory enzyme in the ethylene biosynthetic pathway, were characterized. Genomic clones were analyzed by restriction mapping, and the data in conjunction with sequence comparisons with the previously isolated PCR fragments indicated that at least nine ACC synthase genes (MACS1-9) exist in the banana genome. Southern blot analysis showed they are located in different regions of the banana genome. Three lambda genomic clones (GMACS-1, -9, and -12) were completely sequenced, and gene structures of MACS1 (corresponding to alleles of GMACS-9 and -12) and MACS2 (corresponding to GMACS-1) were elucidated. The coding regions of these three genes were all interrupted by three introns. The size and location of introns are similar to the ACC synthase genes from tomato and Arabidopsis. Northern analysis showed that only MACS1 expressed during fruit ripening and was inducible by exogenous ethylene treatment, which indicates MACS1 is a significant member of the ACC synthase gene family related to ripening in banana fruit. The transcription initiation site of GMACS-12 containing MACS1 was defined. There is a TATTAAT sequence located at position -31 to -25 that qualifies as a TATA box. The delineation of transcription unit in MACS1 will facilitate the promoter studies for this gene and allow its specific functions involved in fruit ripening to be determined.

Molecular cloning and characterization of a novel 1-aminocyclopropane-1-carboxylate oxidase gene involved in ripening of banana fruits.

One novel banana fruit ripening related 1-aminocyclopropane-1-carboxylate (ACC) oxidase gene quite different from ACC oxidase genes from other species was cloned. In contrast to other studies, the polypeptide encoded by this gene, named Mh-ACO1, lacks the putative leucine zipper motif which is conserved in all known ACC oxidases including the other previously reported banana ACC oxidase, Mh-ACO2. The locus consists of two nearly identical paralogous ACC oxidase genes arranged in opposite orientation and separated by a 3.1-kb intergenic region. The has only two introns, at positions identical to , which comprises a coding region interrupted by three introns. The predicted amino acid sequence of Mh-ACO1 shares less than 50% identity to those of ACC oxidase from other climacteric fruits, while that of Mh-ACO2 shows more than 65% homology. When expressed in Saccharomyces cerevisiae -encoded protein possessed the enzyme activity for ethylene conversion. The levels of mRNA corresponding to both and increased during fruit ripening and were induced by exogenous ethylene. We conclude that both and contribute to increased ethylene production in fruits and these two genes are differentially expressed in fruits and other organs in banana.

Type 1 diabetes, cardiovascular complications and sesame ( zhi má).

Diabetes is a major concern among medical practitioners, with the annual mortality rate increasing up to 26.9% in a person aged 65 years or older and 11.3% in the adult. There are many serious complications associated with diabetes, particularly cardiovascular complications due to microvascular diseases. A prerequisite to reduce the risk of microvascular and neurologic complications of type 1 diabetes is normoglycemia. Insulin therapy is the most common treatment used nowadays in type 1 diabetes. However, this method still has many disadvantages such as increased episode of severe hypoglycemia, hypoglycemia unawareness, increased weight gain, transient exacerbation of pre-existing retinopathy, etc. Using insulin pump (the insulin pump is a medical device used for continuous subcutaneous insulin infusion to manage the insulin level in the treatment of diabetes mellitus), is associated with known disadvantages including increased ketoacidosis, infection at the infusion site, and the treatment being less suitable in young children (less than 7 years of age). Therefore, alternative treatment for diabetes is still in great demand. We took the approach of traditional Chinese medicine to discuss this matter. Sesame ( Zhi Má), a herb, has been used medicinally for thousands of years in almost all the countries in the world. The beneficial effects of sesame in remediating diabetes, such as hypoglycemic effects, antioxidant, anti-inflammatory, and hypolipidemic effects, improving fat metabolism, and reducing cholesterol, have been demonstrated in many studies,. However, reports on the effects of sesame in remediating cardiovascular complications in diabetic patients are limited, which necessitates further studies on the effects of sesame on cardiovascular complications.

Expression profiles of a MhCTR1 gene in relation to banana fruit ripening.

The banana (Musa spp.) is a typical climacteric fruit of high economic importance. The development of bananas from maturing to ripening is characterized by increased ethylene production accompanied by a respiration burst. To elucidate the signal transduction pathway involved in the ethylene regulation of banana ripening, a gene homologous to Arabidopsis CTR1 (constitutive triple response 1) was isolated from Musa spp. (Hsien Jin Chiao, AAA group) and designated as MhCTR1. MhCTR1 spans 11.5 kilobases and consists of 15 exons and 14 introns with consensus GT-AG nucleotides situated at their boundaries. MhCTR1 encodes a polypeptide of 805 amino acid residues with a calculated molecular weight of 88.6 kDa. The deduced amino acid sequence of MhCTR1 demonstrates 55%, 56% and 55% homology to AtCTR1, RhCTR1, and LeCTR1, respectively. MhCTR1 is expressed mostly in the mature green pulp and root organs. During fruit development MhCTR1 expression increases just before ethylene production rises. Moreover, MhCTR1 expression was detected mainly in the pulps at ripening stage 3, and correlated with the onset of peel yellowing, while MhCTR1 was constitutively expressed in the peels. MhCTR1 expression could be induced by ethylene treatment (0.01 µL L(-1)), and MhCTR1 expression decreased in both peel and pulp 24 h after treatment. Overall, changes observed in MhCTR1 expression in the pulp closely related to the regulation of the banana ripening process.

Differential expression and functional characterization of the NADPH cytochrome P450 reductase genes from Nothapodytes foetida.

Three unique NADPH:cytochrome P450 reductase (CPR) cDNAs have been isolated from a Nothapodytes foetida cDNA library and characterized. Phylogenetic analysis showed that NfCPR1 is a class I isoform, whereas NfCPR2 and NfCPR3 are class II isoforms. Both NfCPR1 and NfCPR2 transcripts were detected in all examined organs of N. foetida, with the highest level for NfCPR1 being in the seeds whereas for NfCPR2 predominantly in leaves. In contrast, NfCPR3 transcripts were only detected in flower buds and seeds at almost equal expression levels. Moreover, NfCPR1 expression did not change during wounding treatment, whereas NfCPR2 and NfCPR3 were induced in response to wounding. Microsomes isolated from insect cells co-expressing NfCPR2 and cytochrome P450 enzyme geraniol 10-hydroxylase (G10H) enhanced the production of eriodictyol from naringenin approximately 11-fold relative to control G10H-only insect cells, indicating the supportive role of NfCPR2 for G10H activity in insect cells.

Tangy scent in Toona sinensis (Meliaceae) leaflets: isolation, functional characterization, and regulation of TsTPS1 and TsTPS2, two key terpene synthase genes in the biosynthesis of the scent compound.

Toona sinensis (Chinese Mahogany; Meliaceae), a subtropical deciduous tree, has a tangy scent resembling a mix of shallots and garlic. T. sinensis has long been known for its medicinal efficacy for treating enteritis, dysentery, itch and some cancers. However, its volatile components and their biosynthesis remain unexamined. In this study, we identified the spectrum of volatile compounds, isolated and functionally characterized two terpene synthase genes, Tstps1 and Tstps2, responsible for terpenoid synthesis in T. sinensis leaflets. TsTPS1 and TsTPS2 afford multiple products upon incubation with geranyl and farnesyl diphosphate respectively and mainly regulate the biosynthesis of (+) limonene and ß- elemene in vitro, respectively. Headspace analyses show that 98% of leaflet volatiles were sesquiterpenoids and the developing leaflets released a greater diversity and quantity of volatiles than the mature leaflets did, and that ß-elemene was the dominant component in both of them. These data suggested that tangy scent of T. sinensis consists of a combination of terpenoids and that Tstps2 was the major gene involved in the terpenoid biosynthesis in T. sinensis. In situ hybridization revealed that glandular cells of the leaf rachises accumulated abundant Tstps1 mRNA transcripts. Our GFP-based assay further unprecedentedly demonstrated that the transit-peptide of TsTPS1 targets specifically to the mitochondria.

Functional expression of geraniol 10-hydroxylase reveals its dual function in the biosynthesis of terpenoid and phenylpropanoid.

Geraniol 10-hydroxylase (G10H), a cytochrome P450 monooxygenase, has been reported to be involved in the biosynthesis of terpenoid indole alkaloids. The gene for Catharanthus roseus G10H (CrG10H) was cloned and heterologously expressed in baculovirus-infected insect cells. A number of substrates were subjected to assay the enzyme activity of CrG10H. As reported in a previous study, CrG10H hydroxylated the monoterpenoid geraniol at the C-10 position to generate 10-hydroxygeraniol. Interestingly, CrG10H also catalyzed 3'-hydroxylation of naringenin to produce eriodictyol. Coexpression of an Arabidopsis NADPH P450 reductase substantially increased the ability of CrG10H to hydroxylate naringenin. The catalytic activity of CrG10H was approximately 10 times more efficient with geraniol than with naringenin, judged by the k(cat)/K(m) values. Thus, G10H also plays an important role in the biosynthetic pathway of flavonoids, in addition to its previously described role in the metabolism of terpenoids.

cDNA cloning and functional characterization of ETHYLENE INSENSITIVE 3 orthologs from Oncidium Gower Ramsey involved in flower cutting and pollinia cap dislodgement.

The cDNAs encoding ETHYLENE INSENSITIVE3 (EIN3) transcription factor, OgEIL1 and OgEIL2 of Oncidium were cloned, sequenced and characterized. The deduced amino acid sequences of OgEIL1 and OgEIL2 of identified cDNA clones contain all structural features found in the Arabidopsis EIN3, such as an amino terminal acidic domain, a proline-rich region, and five basic conserved domains. Complementation test for OgEIL1 in Arabidopsis ein3 mutant indicate that function of OgEIL1 is the same as Arabidopsis EIN3. RNA gel blot analysis indicated that OgEIL1 and OgEIL2 expressed differentially in the roots, stem, leaves and flower buds of Oncidium. OgEIL1 and OgEIL2 mRNA levels in fully opened flowers increased as time progressed after cutting and reached a maximum in the fifth day and decreased on seventh day, which is consistent with the hypothesis that flowers initiated to wilt when ethylene raised abruptly. In de-capped flowers, OgEIL2 mRNA showed a decrease, while OgEIL1 mRNA exhibited an increase. Exogenous application of ethylene increased the mRNA levels of OgEIL1 and OgEIL2 in flower buds and flowers after cutting compared prior to ethylene treatment, however, in pollinia de-capped flowers, both OgEIL1 and OgEIL2 mRNA levels responded to a decline to exogenous ethylene immediately after treatment. Collectively, it is suggested that the main functions of OgEIL1 and OgEIL2 are to modulate the senescence of Oncidium flowers.

Evaluation of the immunogenicity of a transgenic tobacco plant expressing the recombinant fusion protein of GP5 of porcine reproductive and respiratory syndrome virus and B subunit of Escherichia coli heat-labile enterotoxin in pigs.

Escherichia coli heat-labile enterotoxin B subunit (LTB) can be used as an adjuvant for co-administered antigens. Our previous study showed that the expression of neutralizing epitope GP5 of porcine reproductive and respiratory syndrome virus (PRRSV) in transgenic tobacco plant (GP5-T) could induce PRRSV-specific immune responses in pigs. A transgenic tobacco plant co-expressing LTB and PRRSV GP5 as a fusion protein (LTB-GP5-T) was further constructed and its immunogenicity was evaluated. Pigs were given orally three consecutive doses of equal concentration of recombinant GP5 protein expressed in leaves of LTB-GP5-T or GP5-T at a 2-week interval and challenged with PRRSV at 7 weeks post-initial immunization. Pigs receiving LTB-GP5-T or GP5-T developed PRRSV-specific antibody- and cell-mediated immunity and showed significantly lower viremia and tissue viral load and milder lung lesions than wild type tobacco plant (W-T). The LTB-GP5-T-treated group had relatively higher immune responses than the GP5-T-treated group, although the differences were not statistically significant.