Multifunctional two-dimensional perovskite based solar cells for photodetectors and resistive switching.

The escalating interest in low-dimensional perovskites stems from their tunable optoelectronic traits and robust stability. The pursuit of multifaceted optoelectronic devices holds substantial importance for energy-efficient and space-constrained systems. This investigation showcases the realization of multifunctional two-dimensional perovskite solar cells, incorporating transient light detection and resistive switching functions within a single device, achievable by facile external bias adjustments. Serving as a photodetector, the device exhibits commendable self-powered photodetection attributes, including an exceptionally low dark current density of 1 nA mm-2, a remarkable specific detectivity of 7.67 × 1012 Jones, a swift response time of 0.60 µs, and an expansive linear dynamic range of 72 dB. As a memristor, it showcases enduring performance across 4 × 102 cycles, a substantial on/off ratio of 106, and a rapid operation time of less than 1 µs. This endeavor unveils a pioneering avenue for advancing high-performance, air-stable multifunctional two-dimensional perovskite electronics.

Improved chromosome-level genome assembly of Indian sandalwood (Santalum album).

Santalum album is a well-known aromatic and medicinal plant that is highly valued for the essential oil (EO) extracted from its heartwood. In this study, we present a high-quality chromosome-level genome assembly of S. album after integrating PacBio Sequel, Illumina HiSeq paired-end and high-throughput chromosome conformation capture sequencing technologies. The assembled genome size is 207.39 M with a contig N50 of 7.33 M and scaffold N50 size of 18.31 M. Compared with three previously published sandalwood genomes, the N50 length of the genome assembly was longer. In total, 94.26% of the assembly was assigned to 10 pseudo-chromosomes, and the anchor rate far exceeded that of a recently released value. BUSCO analysis yielded a completeness score of 94.91%. In addition, we predicted 23,283 protein-coding genes, 89.68% of which were functionally annotated. This high-quality genome will provide a foundation for sandalwood functional genomics studies, and also for elucidating the genetic basis of EO biosynthesis in S. album.

Identification of HpMYB1 inducing anthocyanin accumulation in Hippeastrum Hybridum tepals by RNA-seq.

BACKGROUND: Cultivated Hippeastrum × hybridum is a popular ornamental plant with large and colorful flowers, long flowering duration, and high commercial value. As its main ornamental feature, its flower color is related to the anthocyanin content in the tepals. However, the molecular regulatory mechanisms of anthocyanin biosynthesis in H. × hybridum have not yet been elucidated. RESULTS: In the present study, 12 cDNA libraries of four stages of H.× hybridum 'Royal Velvet' tepal development were used for RNA-seq, obtaining 79.83 gigabases (GB) of clean data. The data were assembled into 148,453 unigenes, and 11,262 differentially expressed genes were identified. Forty key enzymes participating in anthocyanin biosynthesis were investigated, and the results showed that most of the anthocyanin structural genes were expressed at low levels in S1 and were markedly upregulated in S2 and S3. The expression profiles of 12 selected genes were verified by qRT-PCR. Furthermore, the R2R3-MYB transcription factor (TF), HpMYB1, involved in the regulation of anthocyanin biosynthesis was identified by sequence, expression pattern, and subcellular localization analyses. Its overexpression in tobacco significantly increased the anthocyanin levels in various tissues and activated anthocyanin-related genes. CONCLUSIONS: Using RNA-seq technology, we successfully identified a potential R2R3-MYB gene, HpMYB1, that regulates anthocyanin biosynthesis in H.× hybridum 'Royal Velvet'. Our findings provide basic transcript information and valuable transcriptome data for further identification of key genes involved in anthocyanin biosynthesis and can be applied in the artificial breeding of new H. × hybridum cultivars with enhanced ornamental value.

Characterization of sandalwood (E,E)-α-farnesene synthase whose overexpression enhances cold tolerance through jasmonic acid biosynthesis and signaling in Arabidopsis.

MAIN CONCLUSION: Santalum album (E,E)-α-farnesene synthase catalyzes FPP into (E,E)-α-farnesene. Overexpression of the SaAFS gene positively improved cold stress tolerance through JA biosynthesis and signaling pathways in Arabidopsis. Volatile terpenoids are released from plants that suffer negative effects following exposure to various biotic and abiotic stresses. Recent studies revealed that (E,E)-α-farnesene synthase (AFS) plays a significant role in a plant's defence against biotic attack. However, little is known about whether AFS contributes to plant resistance to cold stress. In this study, a SaAFS gene was isolated from Indian sandalwood (Santalum album L.) and functionally characterized. The SaAFS protein mainly converts farnesyl diphosphate to (E,E)-α-farnesene. SaAFS was clustered into the AFS clade from angiosperms, suggesting a highly conserved enzyme. SaAFS displayed a significant response to cold stress and methyl jasmonate. SaAFS overexpression (OE) in Arabidopsis enhanced cold tolerance by increasing proline content, reducing malondialdehyde content, electrolyte leakage, and accumulating reactive oxygen species. Transcriptomic analysis revealed that upregulated genes related to stress response and JA biosynthesis and signaling were detected in SaAFS-OE lines compared with wild type plants that were exposed to cold stress. Endogenous JA and jasmonoyl-isoleucine content increased significantly in SaAFS-OE lines exposed to cold stress. Collectively considered, these results suggest that the SaAFS gene is a positive regulator during cold stress tolerance via JA biosynthesis and signaling pathways.

A Cloned Gene HuBADH from Hylocereus undatus Enhanced Salt Stress Tolerance in Transgenic Arabidopsis thaliana Plants.

BACKGROUND: Betaine aldehyde dehydrogenase (BADH) catalyzes the synthesis of glycine betaine and is considered to be a type of osmoregulator, so it can play a role in plants' responses to abiotic stresses. METHODS: In this study, a novel HuBADH gene from Hylocereus undatus (pitaya) was cloned, identified, and sequenced. The full-length cDNA included a 1512 bp open reading frame that encoded a 54.17 kDa protein consisting of 503 amino acids. Four oxidation-related stress-responsive marker genes (FSD1, CSD1, CAT1, and APX2) were analyzed by Quantitative real-time reverse transcription (qRT-PCR) in wild type (WT) and transgenic A. thaiana overexpression lines under NaCl stress. RESULTS: HuBADH showed high homology (79-92%) with BADH of several plants. The HuBADH gene was genetically transformed into Arabidopsis thaliana and overexpressed in transgenic lines, which accumulated less reactive oxygen species than WT plants, and had higher activities of antioxidant enzymes under NaCl stress (i.e., 300 mM). All four marker genes were significantly upregulated in WT and HuBADH-overexpressing transgenic A. thaliana plants under salt stress. Glycine betaine (GB) content was 32-36% higher in transgenic A. thaliana lines than in WT in the control (70-80% in NaCl stress). CONCLUSIONS: Our research indicates that HuBADH in pitaya plays a positive modulatory role when plants are under salt stress.

Transcriptomic analysis reveals biosynthesis genes and transcription factors related to leaf anthocyanin biosynthesis in Aglaonema commutatum.

BACKGROUND: Aglaonema commutatum 'Red Valentine', as a foliage ornamental plant, is widely used for interior and exterior decoration because of its easy cultivation and management. However, reduced proportion of red foliage during large-scale production of A. commutatum seedlings is a frequent occurrence, which has considerable implications on the plant's ornamental and market value. However, the molecular mechanisms underlying this phenomenon remain unclear. RESULTS: To explore the molecular basis of the variation in leaf color of A. commutatum Red Valentine, we performed transcriptome sequencing with the Illumina platform using two different varieties of A. commutatum, namely Red Valentine and a green mutant, at three different stages of leaf development. We annotated 63,621 unigenes and 14,186 differentially expressed genes by pairwise comparison. Furthermore, we identified 26 anthocyanin biosynthesis structural genes. The transcript per million (TPM) values were significantly higher for Red Valentine than for the green mutant in all three developmental stages, consistent with the high anthocyanin content of Red Valentine leaves. We detected positive transcription factors that may be involved in the regulation of anthocyanin biosynthesis using BLAST and through correlation analysis. Downregulation of these transcription factors may downregulate the expression of anthocyanin genes. We obtained full-length cDNA of the anthocyanin biosynthesis and regulatory genes and constructed phylogenetic trees to ensure accuracy of the analysis. CONCLUSIONS: Our study provides insights into the molecular mechanisms underlying leaf variation in A. commutatum Red Valentine and may be used to facilitate the breeding of ornamental cultivars with high anthocyanin levels.

Cloning and functional analysis of 1-deoxy-d-xylulose-5-phosphate synthase (DXS) in Santalum album L.

The commercial value of Santalum album L. lies in its aromatic heartwood and essential oil. Sesquiterpenes are the main components of sandal essential oil, and these are synthesized through the plant's mevalonate (MVA) and methylerythritol phosphate (MEP) pathways. In this study, the first key rate-limiting enzyme, 1-deoxy-d-xylulose-5-phosphate synthase (SaDXS), was investigated to provide a theoretical molecular basis for the sandalwood MEP sesquiterpene biosynthetic pathway. The biofunctions of SaDXS were also analyzed. SaDXS promoters were successfully cloned from a seven-year-old S. album tree. SaDXS1A/1B promoter activity was verified by a ß-glucuronidase (GUS) assay and by analyzing cis-acting elements of the promoters, which carried light- and methyl jasmonate (MeJA)-responsive signals. In an experiment involving yellow S. album seedlings, exposure to light upregulated SaDXS1A/1B expression and increased chlorophyll and carotenoid contents when overexpressed in Arabidopsis thaliana. Analysis of the expression of SaDXS1A/1B and SaSSy, key genes of santalol biosynthesis, revealed SaDXS1A expression in all tissues whereas SaDXS1B was expressed in tissues that contained photosynthetic pigments, such as stems, leaves and flowers. Sandal seedlings exogenously treated with two hormones, MeJA and ethylene, revealed similar expression patterns for SaDXS1A/1B and SaSSy. Sandal seedlings were treated with an inhibitor of DXS, clomazone, but showed no significant changes in the contents of α-santalene, ß-santalene and α-santalol between treatment and control groups. These results suggest that SaDXS1A/1B play a role in the synthesis of sandalwood sesquiterpenes, providing carbon for downstream secondary metabolites. SaDXS1A/1B also play a role in the biosynthesis of chlorophyll, carotenoids, and primary metabolites.

Identification and function of miRNA-mRNA interaction pairs during lateral root development of hemi-parasitic Santalum album L. seedlings.

Sandalwood (Santalum album L.) is a hemi-parasitic tree species famous for its santalol and santalene, which are extracted from its heartwood and roots. The ability to understand root functionality within its branched root system would benefit the regulation of sandalwood growth and enhance the commercial value of sandalwood. Phenotypic and anatomical evidence in this study indicated that seed germination stage 4 (SG4) seemed pivotal for lateral root (LR) morphogenesis. Small RNA (sRNA) high-throughput sequencing of root tissues at three sub-stages of SG4 (lateral root primordia initiation (LRPI), lateral root primordia development (LRPD), and lateral root primordia emergence (LRPE)) was performed to identify microRNAs (miRNAs) associated with LR development. A total of 135 miRNAs, including 70 differentially expressed miRNAs (DEMs), were screened. Ten DEMs were selected to investigate transcript abundance in different organs or developmental stages. Among 100 negative DEM-mRNA interaction pairs, four targets (Sa-miR166m_2, 408d, 858a, and novel_Sa-miR8) were selected for studying cleavage sites by 5' RLM-RACE validation. The expression mode of the four miRNA-mRNA pairs was investigated after indole-3-acetic acid (IAA) treatment. IAA enhanced the abundance of homeobox-leucine-zipper protein 32 (HOX32), laccase 12 (LAC12), myeloblastosis86 (MYB86), and pectin methylesterase inhibitor6 (PMEI6) target transcripts by reducing the expression of Sa-miR166m_2, 408d, 858a, and novel_Sa-miR8 in the first 10 min. A schematic model of miRNA-regulated LR development is proposed for this hemi-parasitic species. This novel genetic information for improving sandalwood root growth and development may allow for the cultivation of fast-growing and high-yielding plantations.

Humic Acid Extracted from Danty via Catalytic Oxidation Using H2O2/Birnessite: Characteristics and Agricultural Beneficial Effects.

Extraction optimization is very important for the quality of humic acid (HA). In this study, actived HA (HAb) was extracted from danty via catalytic oxidation using birnessite as a catalyst and H2O2 as an oxidant. Single-factor experiments and the response surface method were used to optimize the acidic functional group content of HAb. It was found that the maximum acidic functional group content of HAb can be achieved when danty-crushing time, H2O2 concentration, and birnessite dose were 105.7 min, 20, and 2%, respectively. Fourier transform infrared spectra showed that HAb had more surface functional groups than commercial HA (HAc) and HA extracted using the traditional method of the International Humic Substances Society (HAI). In addition, acidic functional group titration showed that HAb had 84.3% more acidic functional groups and 118.9% more carboxyl groups than HAI. Additionally, HAb had the greatest effect on promoting the dissolution of carbonate and bicarbonate, promoting the settlement of calcaline alkaline soil, and improving the germination rate of wheat seeds under saline and alkaline stress. This study provides a basis for the efficient extraction of active HA with rich functional groups and its application in agriculture and many other fields.

Shoot organogenesis and somatic embryogenesis from leaf and petiole explants of endangered Euryodendron excelsum.

Euryodendron excelsum H.T. Chang is a rare and endangered woody plant endemic to China. It is very important to conserve and propagate this species from extinction. In this study, leaves and petioles from the axillary shoots in vitro were used as explants to culture on the different plant growth regulator (PGR) woody plant medium (WPM) and establish an efficient shoot proliferation and plant regeneration system. WPM supplemented with 1.0 mg/L 2,4-D induced callus dedifferentiated into buds and somatic embryos on various media,including PGR-free WPM. However, only adventitious shoots formed on WPM with 1.0 mg/L of cytokinins such as 6-benzyladenine (BA), kinetin (KIN) or thidiazuron (TDZ). When another cytokinin, zeatin, was used, somatic embryos were induced directly from From cut surface of these explants. Adventitious roots could be induced from both explants on WPM with 1.0 mg/L α-naphthaleneacetic acid (NAA). Somatic embryos cultured in PGR-free WPM or WPM with 0.2 mg/L NAA developed roots. Plantlets derived from somatic embryos were transferred to a peat: sand (1:1, v/v) substrate, and showed survival rates of 64.3% at 30 days and 54.6% at 90 days. Callus clumps with adventitious shoot buds that were transferred to WPM containing 1.0 mg/L BA and 0.2 mg/L NAA generated a mean 3.3 multiple shoots. Callus-derived shoots regenerated and rooted successfully (100%) on agar-free vermiculite-based WPM with 0.5 µM NAA after 30 d. Plantlets transplanted to peat soil: vermiculite (1:1, v/v) displayed the highest survival (96.7%) after three months.

AcMYB1 Interacts With AcbHLH1 to Regulate Anthocyanin Biosynthesis in Aglaonema commutatum.

Aglaonema commutatum is one of the most popular foliage plants with abundant leaf phenotypes; therefore, anthocyanin coloration is a vital economic trait in A. commutatum. However, the molecular mechanisms underlying anthocyanin biosynthesis and its regulation remain unclear. In this study, AcMYB1 and AcbHLH1, transcription factor genes related to an R2R3-myeloblast (MYB) and a basic helix-loop-helix (bHLH), respectively, were isolated from A. commutatum "Red Valentine" and functionally characterized. AcMYB1 and AcbHLH1 were found to interact by Y2H and BiFC assay. AcMYB1 was grouped into the AN2 subgroup and shared high homology with the known regulators of anthocyanin biosynthesis. Gene expression analysis showed that both AcMYB1 and AcbHLH1 have similar expression patterns to anthocyanin structural genes and correlate with anthocyanin distribution in different tissues of A. commutatum. Light strongly promoted anthocyanin accumulation by upregulating the expression of anthocyanin-related genes in A. commutatum leaves. Ectopic expression of AcMYB1 in tobacco remarkably increased anthocyanin accumulation in both vegetative and reproductive tissues at various developmental stages. These results provide insights into the regulation of anthocyanin biosynthesis in A. commutatum and are useful for breeding new A. commutatum cultivars with enhanced ornamental value.

Effect of Coated Diammonium Phosphate Combined with Paecilomyces variotii Extracts on Soil Available Nutrients, Photosynthesis-Related Enzyme Activities, Endogenous Hormones, and Maize Yield.

Coated diammonium phosphate (CDAP) is intended to release nutrients steadily in response to the demand of crop growth. A novel biostimulant extracted from Paecilomyces variotii has been shown to regulate gene expression in nutrient transport, enhance nitrogen (N) and phosphorus (P) uptake, and improve nutrient use efficiency. The application of CDAP combined with the Paecilomyces variotii extracts (ZNC) in maize is an efficient approach for reducing waste of resources, improving nutrient supply, and maintaining production stability. The effects of CDAP combined with ZNC on photosynthesis, enzyme activities, endogenous hormone content, maize yield, and P use efficiency (PUE) were investigated in this study. In a pot experiment, CDAP and diammonium phosphate (DAP) were tested together with P levels (1.80, 1.44 g pot-1, P2O5) and two ZNC application rates (0, 4.4 µg pot-1), which included the control treatment that had no P fertilizer added. Results showed that the key influencing elements of maize growth and yield were the soil available-P content, endogenous hormone content, and plant photosynthesis in this study. The combination of DAP and ZNC increased the soil available-P content and the auxin content in leaves at the key stage and hence increased the yield and PUE of maize, compared with DAP. The net photosynthetic rate of CDAP combined with ZNC was higher by 23.1% than that of CDAP alone, as well as by 32.0% than that of DAP combined with ZNC. Moreover, the combination of CDAP and ZNC increased the yield and PUE by 8.2% and 15.6 percentage points compared with DAP combined with ZNC while increasing the yield and PUE compared with CDAP. In conclusion, combining CDAP with ZNC as an environmentally friendly fertilizer could improve photosynthesis-related enzyme activity and enhance the net photosynthetic rate, resulting in an increase in maize yield and PUE significantly.

Exogenous GA3 promotes flowering in Paphiopedilum callosum (Orchidaceae) through bolting and lateral flower development regulation.

Paphiopedilum orchids have a high ornamental value, and their flower abundance and timing are both key horticultural traits regulated by phytohormones. All one-flowered Paphiopedilum have additional lateral buds in the apical bract that fail to develop. In this study, an exogenous gibberellin (GA3) application promoted flowering of Pathiopedilum callosum by inducing its early bolting instead of the floral transition of dominant flowers. Applying GA3 effectively promoted lateral flower differentiation, resulting in a two-flowered inflorescence. GA-promoted lateral flower formation involved GA interacting with indole-3-acetic acid (IAA) and cytokinins (CTKs), given the decreased CTK content and downregulated expression of CTK synthesis genes, the increased IAA content and downregulated expression of IAA degradation, and the upregulated expression of transport genes. Further, GA acted via PcDELLA, PcTCP15, and PcXTH9 expressed in stage 5 to promote bolting, and via expression of PcAP3, PcPI, and PcSEP to promote flowering. This study provides insight into mechanisms regulating flower development of P. callosum.

Identification and Functional Analysis of SabHLHs in Santalum album L.

Santalum album L., a semi-parasitic evergreen tree, contains economically important essential oil, rich in sesquiterpenoids, such as (Z) α- and (Z) ß-santalol. However, their transcriptional regulations are not clear. Several studies of other plants have shown that basic-helix-loop-helix (bHLH) transcription factors (TFs) were involved in participating in the biosynthesis of sesquiterpene synthase genes. Herein, bHLH TF genes with similar expression patterns and high expression levels were screened by co-expression analysis, and their full-length ORFs were obtained. These bHLH TFs were named SaMYC1, SaMYC3, SaMYC4, SaMYC5, SabHLH1, SabHLH2, SabHLH3, and SabHLH4. All eight TFs had highly conserved bHLH domains and SaMYC1, SaMYC3, SaMYC4, and SaMYC5, also had highly conserved MYC domains. It was indicated that the eight genes belonged to six subfamilies of the bHLH TF family. Among them, SaMYC1 was found in both the nucleus and the cytoplasm, while SaMYC4 was only localized in the cytoplasm and the remaining six TFs were localized in nucleus. In a yeast one-hybrid experiment, we constructed decoy vectors pAbAi-SSy1G-box, pAbAi-CYP2G-box, pAbAi-CYP3G-box, and pAbAi-CYP4G-box, which had been transformed into yeast. We also constructed pGADT7-SaMYC1 and pGADT7-SabHLH1 capture vectors and transformed them into bait strains. Our results showed that SaMYC1 could bind to the G-box of SaSSy, and the SaCYP736A167 promoter, which SaSSy proved has acted as a key enzyme in the synthesis of santalol sesquiterpenes and SaCYP450 catalyzed the ligation of santalol sesquiterpenes into terpene. We have also constructed pGreenII 62-SK-SaMYC1, pGreenII 0800-LUC-SaSSy and pGreenII 0800-LUC-SaCYP736A167 via dual-luciferase fusion expression vectors and transformed them into Nicotiana benthamiana using an Agrobacterium-mediated method. The results showed that SaMYC1 was successfully combined with SaSSy or SaCYP736A167 promoter and the LUC/REN value was 1.85- or 1.55-fold higher, respectively, than that of the control group. Therefore, we inferred that SaMYC1 could activate both SaSSy and SaCYP736A167 promoters.

Genome-Wide Identification and Analysis of NAC Transcription Factor Family in Two Diploid Wild Relatives of Cultivated Sweet Potato Uncovers Potential NAC Genes Related to Drought Tolerance.

NAC (NAM, ATAF1/2, and CUC2) proteins play a pivotal role in modulating plant development and offer protection against biotic and abiotic stresses. Until now, no systematic knowledge of NAC family genes is available for the food security crop, sweet potato. Here, a comprehensive genome-wide survey of NAC domain-containing proteins identified 130 ItbNAC and 144 ItfNAC genes with full length sequences in the genomes of two diploid wild relatives of cultivated sweet potato, Ipomoea triloba and Ipomoea trifida, respectively. These genes were physically mapped onto 15 I. triloba and 16 I. trifida chromosomes, respectively. Phylogenetic analysis divided all 274 NAC proteins into 20 subgroups together with NAC transcription factors (TFs) from Arabidopsis. There were 9 and 15 tandem duplication events in the I. triloba and I. trifida genomes, respectively, indicating an important role of tandem duplication in sweet potato gene expansion and evolution. Moreover, synteny analysis suggested that most NAC genes in the two diploid sweet potato species had a similar origin and evolutionary process. Gene expression patterns based on RNA-Seq data in different tissues and in response to various hormone, biotic or abiotic treatments revealed their possible involvement in organ development and response to various biotic/abiotic stresses. The expression of 36 NAC TFs, which were upregulated in the five tissues and in response to mannitol treatment, was also determined by real-time quantitative polymerase chain reaction (RT-qPCR) in hexaploid cultivated sweet potato exposed to drought stress. Those results largely corroborated the expression profile of mannitol treatment uncovered by the RNA-Seq data. Some significantly up-regulated genes related to drought stress, such as ItbNAC110, ItbNAC114, ItfNAC15, ItfNAC28, and especially ItfNAC62, which had a conservative spatial conformation with a closely related paralogous gene, ANAC019, may be potential candidate genes for a sweet potato drought tolerance breeding program. This analysis provides comprehensive and systematic information about NAC family genes in two diploid wild relatives of cultivated sweet potato, and will provide a blueprint for their functional characterization and exploitation to improve the tolerance of sweet potato to abiotic stresses.

Characterization of phytohormone and transcriptome profiles during protocorm-like bodies development of Paphiopedilum.

BACKGROUND: Paphiopedilum, commonly known as slipper orchid, is an important genus of orchid family with prominent horticultural value. Compared with conventional methods such as tillers and in vitro shoots multiplication, induction and regeneration of protocorm-like bodies (PLBs) is an effective micropropagation method in Paphiopedilum. The PLB initiation efficiency varies among species, hybrids and varieties, which leads to only a few Paphiopedilum species can be large-scale propagated through PLBs. So far, little is known about the mechanisms behind the initiation and maintenance of PLB in Paphiopedilum. RESULTS: A protocol to induce PLB development from seed-derived protocorms of Paphiopedilum SCBG Huihuang90 (P. SCBG Prince × P. SCBG Miracle) was established. The morphological characterization of four key PLB developmental stages showed that significant polarity and cell size gradients were observed within each PLB. The endogenous hormone level was evaluated. The increase in the levels of indoleacetic acid (IAA) and jasmonic acid (JA) accompanying the PLBs differentiation, suggesting auxin and JA levels were correlated with PLB development. Gibberellic acid (GA) decreased to a very low level, indicated that GA inactivation may be necessary for shoot apical meristem (SAM) development. Comparative transcriptomic profiles of four different developmental stages of P. SCBG Huihuang90 PLBs explore key genes involved in PLB development. The numbers of differentially expressed genes (DEGs) in three pairwise comparisons (A vs B, B vs C, C vs D) were 1455, 349, and 3529, respectively. KEGG enrichment analysis revealed that DEGs were implicated in secondary metabolite metabolism and photosynthesis. DEGs related to hormone metabolism and signaling, somatic embryogenesis, shoot development and photosynthesis were discussed in detail. CONCLUSION: This study is the first report on PLB development in Paphiopedilum using transcriptome sequencing, which provides useful information to understand the mechanisms of PLB development.

Characterization of embryo and protocorm development of Paphiopedilum spicerianum.

Paphiopedilum spicerianum (P. spicerianum) is a rare orchid species with high ornamental value. Asymbiotic germination is the most efficient propagation method for conservation and commercial purposes because clonal propagation is very difficult and the separation of native species of Paphiopedilum through aseptic seeding is uncommon owing to their conservatism. However, a high protocorm developmental arresting rate during the asymbiotic germination is the major obstacle for seedling establishment. The fundamental understanding of embryo and protocorm developmental mechanisms will guide the development of an effective propagation method. The morphological and physiological characterization of the key developmental process of embryos and protocorms shows that the mature seeds of P. spicerianum consist of a spherical embryo without an endosperm. Seed coats become heavily lignified once the embryo is mature. Embryo cell size is relatively uniform, and significant structure polarity and cell size gradients occur at the early protocorm stage. The high level of auxin and cytokinin accumulation at the early stage of embryo development and protocorm stage may help to facilitate cell division. The transcriptome profiles of protocorms at three different developmental stages were compared to explore the regulatory mechanism of protocorm development. Kyoto Encyclopedia of Genes and Genomes enrichment analysis revealed that differentially expressed genes were implicated in secondary metabolite metabolism, plant hormone signal transduction and photosynthesis. The temporal expression patterns of candidate genes related to embryo and shoot development were analyzed to reveal their roles in protocorm development: in the early stage of protocorm development, embryonic development related genes such as SERKs and BBM1 were active, while in the late stage of protocorm, shoot apical meristem related genes such as WOX8, CLAVATA2, CUC2, and SCR were active.

Cloning and expression analysis of mevalonate kinase and phosphomevalonate kinase genes associated with the MVA pathway in Santalum album.

Sandalwood (Santalum album L.) is highly valued for its fragrant heartwood and extracted oil. Santalols, which are the main components of that oil, are terpenoids, and these are biosynthesized via the mevalonic acid (MVA) pathway. Mevalonate kinase (MK) and phosphomevalonate kinase (PMK) are key enzymes in the MVA pathway. Little is known about the genes that encode MK and PMK in S. album or the mechanism that regulates their expression. To isolate and identify the functional genes involved in santalol biosynthesis in S. album, an MK gene designated as SaMK, and a PMK gene designated as SaPMK, were cloned from S. album. The sequences of these genes were analyzed. A bioinformatics analysis was conducted to assess the homology of SaMK and SaPMK with MK and PMK genes from other plants. The subcellular localization of SaMK and SaPMK proteins was also investigated, as was the functional complementation of SaMK and SaPMK in yeast. Our results show that the full-length cDNA sequences of SaMK and SaPMK were 1409 bp and 1679 bp long, respectively. SaMK contained a 1381 bp open reading frame (ORF) encoding a polypeptide of 460 amino acids and SaPMK contained a 1527 bp ORF encoding a polypeptide of 508 amino acids. SaMK and SaPMK showed high homology with MK and PMK genes of other plant species. Functional complementation of SaMK in a MK-deficient mutant yeast strain YMR208W and SaPMK in a PMK-deficient mutant yeast strain YMR220W confirmed that cloned SaMK and SaPMK cDNA encode a functional MK and PMK, respectively, mediating MVA biosynthesis in yeast. An analysis of tissue expression patterns revealed that SaMK and SaPMK were constitutively expressed in all the tested tissues. SaMK was highly expressed in young leaves but weakly expressed in sapwood. SaPMK was highly expressed in roots and mature leaves, but weakly expressed in young leaves. Induction experiments with several elicitors showed that SaMK and SaPMK expression was upregulated by methyl jasmonate. These results will help to further study the role of MK and PMK genes during santalol biosynthesis in S. album.

Molecular Cloning and Functional Analysis of 1-Deoxy-D-Xylulose 5-Phosphate Reductoisomerase from Santalum album.

Sandalwood (Santalum album L.) heartwood-derived essential oil contains a high content of sesquiterpenoids that are economically highly valued and widely used in the fragrance industry. Sesquiterpenoids are biosynthesized via the mevalonate acid and methylerythritol phosphate (MEP) pathways, which are also the sources of precursors for photosynthetic pigments. 1-deoxy-D-xylulose-5-phosphate reductoisomerase (DXR) is a secondary rate-limiting enzyme in the MEP pathway. In this paper, the 1416-bp open reading frame of SaDXR and its 897-bp promoter region, which contains putative conserved cis-elements involved in stress responsiveness (HSE and TC-rich repeats), hormone signaling (abscisic acid, gibberellin and salicylic acid) and light responsiveness, were cloned from 7-year-old S. album trees. A bioinformatics analysis suggested that SaDXR encodes a functional and conserved DXR protein. SaDXR was widely expressed in multiple tissues, including roots, twigs, stem sapwood, leaves, flowers, fruit and stem heartwood, displaying significantly higher levels in tissues with photosynthetic pigments, like twigs, leaves and flowers. SaDXR mRNA expression increased in etiolated seedlings exposed to light, and the content of chlorophylls and carotenoids was enhanced in all 35S::SaDXR transgenic Arabidopsis thaliana lines, consistent with the SaDXR expression level. SaDXR was also stimulated by MeJA and H2O2 in seedling roots. α-Santalol content decreased in response to fosmidomycin, a DXR inhibitor. These results suggest that SaDXR plays an important role in the biosynthesis of photosynthetic pigments, shifting the flux to sandalwood-specific sesquiterpenoids.

Elicitors Modulate Young Sandalwood (Santalum album L.) Growth, Heartwood Formation, and Concrete Oil Synthesis.

Five chemical elicitors--6-benzyladenine (BA), ethephon (ETH), methyl jasmonate (MeJA), hydrogen peroxide (H2O2) and calcium chloride (CaCl2)--were used to treat 1- and 5-year-old sandal trees (Santalum album L.) to assess their effects on growth, heartwood formation and concrete oil synthesis. The results showed that some newly formed branches in stems that were induced by BA and ETH displayed leaf senescence and developed new smaller and light-green leaves. The relative percentage of concrete oil from the heartwood of water-treated trees (0.65%) was significantly lower than that from trees treated with 4 mM H2O2 (2.85%) and 4 mM BA (2.75%) within one year. Four mM BA, H2O2 and CaCl2 induced a significantly higher level of sesquiterpenoids than heartwood treated with 2 mM of these elicitors. Four mM MeJA induced significantly less sesquiterpenoids than heartwood treated with 2 mM MeJA. Morphological, physiological, and chromatographic-spectrometric technologies were integrated to trace the potential function of these exogenously applied chemical elicitors. The results may have important applications and provide a better understanding of the molecular mechanism of heartwood formation and hardening in young sandalwood trees.