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Though many biological roles of ethylene have been investigated intensively, the molecular mechanism of ethylene's action in woody plants remains unclear. In this study, we investigated the effects of exogenous 1-aminocyclopropane-1-carboxylic acid (ACC), the precursor of ethylene, on the growth of Neolamarckia cadamba seedlings, a fast-growing tropical tree. After 14 days of ACC treatment, the plants showed a reduced physiological morphology while stem diameter increased; however, this did not occur after the addition of 1-MCP. Meanwhile, the lignin content of N. cadamba also increased. Transcriptome analysis revealed that the expression of the ethylene biosynthesis and signaling genes ACC oxidase (ACO) and ethylene insensitive 3 (EIN3) were up-regulated mainly at the 6th hour and the 3rd day of the ACC treatment, respectively. The transcription levels of transcription factors, mainly in the basic helix-loop-helix (bHLH), ethylene response factor (ERF), WRKY and v-myb avian myeloblastosis viral oncogene homolog (MYB) families, involved in the ethylene signaling and secondary growth also increased significantly. Furthermore, in accordance to the increased lignification of the stem, the transcriptional level of key enzymes in the phenylalanine pathway were elevated after the ACC treatment. Our results revealed the physiological and molecular mechanisms underlying the secondary growth stimulated by exogenous ACC treatment on N. cadamba seedlings.

Neolamarckia cadamba (N. cadamba) is a fast-growing tree species with tremendous economic and ecological value; the study of the key genes regulating photosynthesis and sugar accumulation is very important for the breeding of N. cadamba. Fructose 1,6-bisphosptase (FBP) gene has been found to play a key role in plant photosynthesis, sugar accumulation and other growth processes. However, no systemic analysis of FBPs has been reported in N. cadamba. A total of six FBP genes were identifed and cloned based on the N. cadamba genome, and these FBP genes were sorted into four groups. The characteristics of the NcFBP gene family were analyzed such as phylogenetic relationships, gene structures, conserved motifs, and expression patterns. A cis-acting element related to circadian control was first found in the promoter region of FBP gene. Phylogenetic and quantitative real-time PCR analyses showed that NcFBP5 and NcFBP6 may be chloroplast type 1 FBP and cytoplasmic FBP, respectively. FBP proteins from N. cadamba and 22 other plant species were used for phylogenetic analyses, indicating that FBP family may have expanded during the evolution of algae to mosses and differentiated cpFBPase1 proteins in mosses. This work analyzes the internal relationship between the evolution and expression of the six NcFBPs, providing a scientific basis for the evolutionary pattern of plant FBPs, and promoting the functional studies of FBP genes.

Artificial induction of polyploidy is widely used in breeding programmes to improve the agronomic traits. The drumstick tree (Moringa oleifera Lam.) has a range of potential commercial uses, as the vegetative organs have high nutritional, medicinal, and feed values. In the present study, in vitro tetraploidisation in drumstick tree was performed by treating leaf segments with colchicine and subsequently verifying the ploidy levels. For polyploidisation, explant survival and regeneration rates were affected more by exposure time than by colchicine concentration, and the highest polyploidisation efficiency was observed at 500 mg/L colchicine for 3 days, which yielded 21% tetraploids. The morphological characteristics and contents of seven fodder-related nutrients (crude protein, ether extract, ash, acid detergent fibre, neutral detergent fibre, calcium, and phosphorus) in the leaves and shoots were compared between tetraploid and diploid drumstick trees. The resulting tetraploids showed significantly enhanced leaf and stomatal size. In addition, the contents of seven fodder-related nutrients were higher, although to varying degrees, in tetraploids than in diploids. The results indicated that the tetraploid produced in this study exhibited superior agronomical traits and improved biomass yield than diploids, and may represent excellent raw materials for fodder to enhance biomass and nutrition.

Neolamarckia cadamba (Roxb.) Bosser is a tropical evergreen broadleaf tree species that could play an important role in meeting the increasing demand for wood products. However, multi-level genetic variation and selection efficiency for growth traits in N. cadamba is poorly characterized. We therefore investigated the efficiency of early selection in N. cadamba by monitoring the height (HT), diameter at breast height (DBH), and tree volume (V) in 39 half-sib families from 11 provenances at ages 2, 3, 4, 5, and 6 years in a progeny test. Age-related trends in growth rate, genetic parameters in multi-level, efficiency of early selection, and realized gain in multi-level for growth traits were analyzed. The result showed that genetic variation among families within provenances was higher than that among provenances. The estimated individual heritability values for the growth traits ranged from 0.05 to 0.26, indicating that the variation of growth traits in N. cadamba was subject to weak or intermediate genetic control. The age–age genetic correlations for growth traits were always positive and high (0.51–0.99), and the relationships between the genetic/phenotypic correlations and the logarithm of the age ratio (LAR) were described well by linear models (R2 &gt, 0.85, except the fitting coefficient of genetic correlation and LAR for HT was 0.35). On the basis of an early selection efficiency analysis, we found that it is the best time to perform early selection for N. cadamba at age 5 before half-rotation, and the selection efficiencies were 157.28%, 151.56%, and 127.08% for V, DBH, and HT, respectively. Higher realized gain can be obtained by selecting superior trees from superior families. These results can be expected to provide theoretical guidance and materials for breeding programs in N. cadamba and can even be a reference for breeding strategies of other fast-growing tree species.

Neolamarckia cadamba is an important fast growing tree species used for pulping and wood material in industry for it’s desirable wood properties. As one of the most important content in wood, lignin provides structural integrity, strength, and hydrophobicity to the thickened cell walls and is the major factor contributing to biomass recalcitrance. It does not reduce the palatability of forage grass for animals, but it hinders the isolation of cellulose fibers and the efficient enzymatic depolymerization of cellulose and hemicellulose into fermentable sugars in biorefining processes by limiting the access by hydrolytic enzymes to their polysaccharide substrates. This work focused on analyzing the functions of NcCSE (Caffeoyl Shikimate Esterase, GenBank accession number: MG739672) and NcHCT (Hydroxycinnamoyl Transferase,GenBank accession number: MG739673) in the lignin biosynthetic process in order to improve the potential for utilization of leaves and wood from N. cadamba. The mutant phenotype of cse-2 was dramatically complemented to WT in the stable transgenic lines cse-35S::NcCSE, but overexpression of NcHCT in the cse-2 mutant did not have the same result as cse-35S::NcCSE, providing only partial complementation.

Moringa oleifera Lam. (MO) is a fast-growing multi-purpose deciduous tree with high biomass and nutritional value. However, the presence of antinutritional factors, poor palatability, and indigestibility of Moringa oleifera leaf meal (MOLM) restrict its application to animal feed. This study aimed to obtain high-quality protein feeds via solid-state fermentation (SSF) of MOLM. The process conditions for increasing the true protein (TP) content using Aspergillus niger, Candida utilis and Bacillus subtilis co-cultures were optimized, and the chemical composition of MOLM was compared before and after fermentation. The results of this study showed that the highest TP content could be obtained through mixed-strain culture of A. niger, C. utilis and B. subtilis at a ratio of 1:1:2. The MOLM was inoculated with A. niger, followed by C. utilis and B. subtilis 24 h later. The optimized co-culture parameters were as follows: total inoculation size, 24%; temperature, 32 °C; fermentation time, 6.5 days; and initial water content, 60%. The maximum TP yield was 28.37%. Notably, in the fermented MOLM (FMOLM), the content of nutrients such as crude protein (CP), small peptides, and total amino acids (AAs) were significantly increased relative to unfermented MOLM, whereas the contents of crude fiber (CF), tannin, and phytic acid were significantly decreased. MOLM analysis using scanning electron microscopy (SEM) revealed that SSF disrupted the surface structure of MOLM, and sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) indicated that macromolecular proteins were degraded. The in vitro protein digestibility (IVPD) of FMOLM was also improved significantly. Our findings suggest that multi-strain fermentation with A. niger, C. utilis and B. subtilis improves the nutritional quality of MOLM, rendering it a viable functional feedstuff for use in livestock industries in the future.

To study the effects of citric acid on fermentation process of Amomum villosum silage, A. villosum was ensiled without or with 1%, 2% citric acid and fermentation parameters and bacterial diversity were analyzed after 3, 7, 14, 30 days ensiling, respectively. Citric acid treated silages had lower dry matter loss (1.83% vs 2.23%), pH (3.84 vs 6.02), ammonia-N (0.33 vs 1.79 g/kg DM) and coliform bacteria number (2.00 vs 7.27 log

Greenhouse gas emissions from ruminants, including methane (CH4) and carbon dioxide (CO2), make a significant contribution to global warming. Exploring novel and effective approaches for reducing ruminant greenhouse gas emissions is essential for cleaner livestock production and alleviating global warming. To investigate the effectiveness of a new feed source, Neolamarckia cadamba leaves (NCL), in reducing greenhouse gas emissions from ruminants as well as the underlying mechanism, 0%, 10%, 30%, and 50% NCL were ensiled with corn stalk in two trials. The results indicated that NCL inclusion increased the crude protein content (P

A better understanding of biomass usability during storage would offer basis for management decisions in production. High-moisture corn stover was ensiled with sulfuric acid (H2SO4, 0.3% and 0.6%) or sodium hydroxide (NaOH, 0.5% and 1.0%) and ensiling characteristics, lignocellulosic profile and enzymatic saccharification were investigated on day 3, 7, 15, 30 and 60 of ensiling. The results showed that 0.6% H2SO4 reduced dry matter loss (9.81% to 6.34%) and ammonia-N content (3.89 to 1.04 g/kg DM) during ensiling, whereas it was converse for NaOH treatment (19.89%, 5.74 g/kg DM). Hemicellulose was reduced (27.98% to 22.61%, 16.81% DM) by 0.6% H2SO4 or 1.0% NaOH. Saccharification yield was decreased (306 to 229 mg/g DM) during ensiling, which was improved (229 to 356, 277 mg/g DM) by H2SO4 and NaOH treatments. This study suggests that ensiling with addition of 0.6% H2SO4 could improve nutrient preservation and saccharification yield of high-moisture corn stover.

To investigate the effects of wilting and lactic acid bacterial inoculants on the fermentation quality and bacterial community of Moringa oleifera leaf silage, fresh and wilted M. oleifera leaves were ensiled with or without Lactobacillus farciminis LF or Lactococcus lactis LL for 1, 7, 14, 30, and 60 days. The results showed that wilting, inoculants, and their interaction exerted significant (P

WRKY proteins belong to one of the largest families of transcription factors. They have important functions in plant growth and development, signal transduction and stress responses. However, little information is available regarding the WRKY family in drumstick (Moringa oleifera Lam.). In the present study, we identified 54 MoWRKY genes in this species using genomic data. On the basis of structural features of the proteins they encode, the MoWRKY genes were classified into three main groups, with the second group being further divided into five subgroups. Phylogenetic trees constructed from the sequences of WRKY domains and overall amino acid compositions derived from drumstick and Arabidopsis were similar; the results indicated that the WRKY domain was the main evolutionary unit of WRKY genes. Gene structure and conserved motif analysis showed that genes with similar structures and proteins with similar motif compositions were usually clustered in the same class. Selective pressure analysis indicated that although neutral evolution and positive selection have happened in several MoWRKY genes, most have evolved under strong purifying selection. Moreover, different subgroups had evolved at different rates. The levels of expression of MoWRKY genes in response to five different abiotic stresses (salt, heat, drought, H2O2, cold) were evaluated by reverse transcription polymerase chain reaction (RT-PCR) and quantitative RT-PCR (qRT-PCR), with the results indicating that these genes had different expression levels and that some may be involved in abiotic stress responses. Our results will provide a foundation for cloning genes with specific functions for use in further research and applications.

Four lactic acid bacteria strains (LP, LF, LL, W), isolated and selected from Moringa oleifera leaves (MOL) silage, were identified as Lactobacillus plantarum, L. farciminis, Lactococcus lactis, Weissella thailandensis, respectively. Fermentation quality and bacterial community of MOL ensiled without or with the four strains at 15 °C and 30 °C were investigated. Results showed that all the LAB strains decreased (P 0.05) the pH and ammonia-N content of MOL silage. Silage stored at 30 °C showed higher (P 0.05) DM loss, acetic acid and ammonia-N content, and lower LAB count than 15 °C. The environmental temperature also made a great influence on bacterial community of MOL silage. Bacterial diversity was lower and the abundance of Lactobacillus was higher in silages stored at 15 °C. In conclusion, LAB inoculants and a relatively low environmental temperature could be effective to improve the quality of MOL silage.

The goal of this paper was to investigate the effects of polyphenol oxidases (PPO) on proteolysis and lipolysis during ensiling of Moringa oleifera leaves with or without pyrocatechol. Directly, 30 U/g and 60 U/g PPO with or without 1% pyrocatechol were added to M.oleifera leaves. Fermentation characteristic, microbial population, antioxidant capacity, protein fraction and lipids composition were analyzed after 10 days and 20 days ensiling. The addition of PPO alone showed little influence on the silage quality, though it decreased (p

Mixed ensiling is believed an effective way to improve nutrient preservation and utilization. The effect of mixing Moringa oleifera leaves (MOL) on silage quality, aerobic stability and microbial communities of rice straw ensiled with/without propionic acid were investigated after 140 days fermentation. The results showed that mixing MOL decreased the pH (4.69 vs 3.85), butyric acid (17.4 g/kg DM vs not detected), ammonia-N (3.36 vs 2.17 g/kg DM) and fiber contents (626 vs 462 g/kg DM) but increased protein content (88.4 vs 125 g/kg DM) of rice straw silages. It also increased the relative abundance of Lactobacillus (12.96% vs 50.82%) at unsealing and Issatchenkia (2.02% vs 36.03%) after exposure to air, and might improve aerobic stability of silages. The addition of propionic acid could inhibit deaminization activity during ensiling and exposure to air. It is suggested mixing MOL could improve the fermentation quality and nutrition of rice straw silage.

Bauhinia variegate flower (BVF) was supposed to improve silage fermentation due to its abundant active components. Thus, corn stalk and stylo were ensiled with addition of 0, 5% or 10% BVF, and then ensiling characteristics, protein fraction and bacterial community were analyzed after 60-day fermentation. The contents of butyric acid (2.9 vs not detected, 13.2 vs 3.0 g/kg DM in corn stalk and stylo silage, respectively), ammonia-N (100.2 vs 83.2, 110.8 vs 61.9 g/kg total N) and free amino acid (35.6 vs 16.5, 35.0 vs 16.4 g/kg total N) were decreased in 10% BVF treated silages. The bacterial diversity was increased, where the relative abundance of Enterobacter or Clostridium decreased and that of lactic acid producing bacteria such as Lactobacillus, Weissella or Enterococcus increased. It is suggested that BVF could be used to improve fermentation quality and nutrient preservation of high-moisture corn stalk and stylo silage.

WRKY proteins belong to one of the largest families of transcription factors. They have important functions in plant growth and development, signal transduction and stress responses. However, little information is available regarding the WRKY family in drumstick (Moringa oleifera Lam.). In the present study, we identified 54 MoWRKY genes in this species using genomic data. On the basis of structural features of the proteins they encode, the MoWRKY genes were classified into three main groups, with the second group being further divided into five subgroups. Phylogenetic trees constructed from the sequences of WRKY domains and overall amino acid compositions derived from drumstick and Arabidopsis were similar; the results indicated that the WRKY domain was the main evolutionary unit of WRKY genes. Gene structure and conserved motif analysis showed that genes with similar structures and proteins with similar motif compositions were usually clustered in the same class. Selective pressure analysis indicated that although neutral evolution and positive selection have happened in several MoWRKY genes, most have evolved under strong purifying selection. Moreover, different subgroups had evolved at different rates. The levels of expression of MoWRKY genes in response to five different abiotic stresses (salt, heat, drought, H2O2, cold) were evaluated by reverse transcription polymerase chain reaction (RT-PCR) and quantitative RT-PCR (qRT-PCR), with the results indicating that these genes had different expression levels and that some may be involved in abiotic stress responses. Our results will provide a foundation for cloning genes with specific functions for use in further research and applications.

To investigate the reason for well preservation of protein in Neolamarckia cadamba leaves (NCL) during ensiling, fresh NCL were ensiled with or without addition of 2.0 mL/kg formic acid (FA) or 1.0 × 109 CFU/kg Lactobacillus farciminis (LF), and the dynamics of protease activity and microbial community were analyzed. Nonprotein-N, free amino acid, ammonia-N, the activities of carboxypeptidase and aminopeptidase, and bacterial diversity were low during NCL ensiling. Exiguobacterium dominated in NCL silage and its relative abundance increased while Enterobacter abundance decreased during ensiling. FA lowered (P

Sequence-related amplified polymorphism (SRAP) markers were used to investigate the genetic diversity among 30 populations of Toona ciliata Roem. sampled from the species’ distribution area in China. To analyze the polymorphism in the SRAP profiles, 1505 primer pairs were screened and 24 selected. A total of 656 SRAP bands ranging from 100 to 1500 bp were acquired, of these 505 bands (77%) were polymorphic. The polymorphism information content (PIC) values ranged from 0.32 to 0.45, with an average of 0.41. An analysis of molecular variance (AMOVA) indicated that the most significant variation was attributable to differences among the populations and that variation within the populations was small. STRUCTURE analysis divided the 30 populations into two parts. The unweighted pair group method of arithmetic averages (UPGMA) clustering and principal coordinates analysis (PCoA) showed that the 30 populations could be classified into four types. The results demonstrate a clear geographical trend for T. ciliata in China and provide a theoretical basis for future breeding and conservation strategy of T. ciliata.

The uncertainty about whether, in China, the genus Melia (Meliaceae) consists of one species (M. azedarach Linnaeus) or two species (M. azedarach and M. toosendan Siebold & Zuccarini) remains to be clarified. Although the two putative species are morphologically distinguishable, genetic evidence supporting their taxonomic separation is lacking. Here, we investigated the genetic diversity and population structure of 31 Melia populations across the natural distribution range of the genus in China. We used sequence-related amplified polymorphism (SRAP) markers and obtained 257 clearly defined bands amplified by 20 primers from 461 individuals. The polymorphic loci (P) varied from 35.17% to 76.55%, with an overall mean of 58.24%. Nei’s gene diversity (H) ranged from 0.13 to 0.31, with an overall mean of 0.20. Shannon’s information index (I) ranged from 0.18 to 0.45, with an average of 0.30. The genetic diversity of the total population (Ht) and within populations (Hs) was 0.37 ± 0.01 and 0.20 ± 0.01, respectively. Population differentiation was substantial (Gst = 0.45), and gene flow was low. Of the total variation, 31.41% was explained by differences among putative species, 19.17% among populations within putative species, and 49.42% within populations. Our results support the division of genus Melia into two species, which is consistent with the classification based on the morphological differentiation.