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Carotenoid cleavage oxygenases can cleave carotenoids into a range of biologically important products. Carotenoid isomerooxygenase (NinaB) and β, β-carotene 15, 15\'-monooxygenase (BCO1) are two important oxygenases. In order to understand the roles that both oxygenases exert in crustaceans, we first investigated NinaB-like (EsNinaBl) and BCO1-like (EsBCO1l) within the genome of Chinese mitten crab (Eriocheir sinensis). Their functions were then deciphered through an analysis of their expression patterns, an in vitro β-carotene degradation assay, and RNA interference. The results showed that both EsNinaBl and EsBCO1l contain an RPE65 domain and exhibit high levels of expression in the hepatopancreas. During the molting stage, EsNinaBl exhibited significant upregulation in stage C, whereas EsBCO1l showed significantly higher expression levels at stage AB. Moreover, dietary supplementation with β-carotene resulted in a notable increase in the expression of EsNinaBl and EsBCO1l in the hepatopancreas. Further functional assays showed that the EsNinaBl expressed in E. coli underwent significant changes in its color, from orange to light; in addition, its β-carotene cleavage was higher than that of EsBCO1l. After the knockdown of EsNinaBl or EsBCO1l in juvenile E. sinensis, the expression levels of both genes were significantly decreased in the hepatopancreas, accompanied by a notable increase in the redness (a*) values. Furthermore, a significant increase in the β-carotene content was observed in the hepatopancreas when EsNinaBl-mRNA was suppressed, which suggests that EsNinaBl plays an important role in carotenoid cleavage, specifically β-carotene. In conclusion, our findings suggest that EsNinaBl and EsBCO1l may exhibit functional co-expression and play a crucial role in carotenoid cleavage in crabs.

BACKGROUND: The domain of unknown function 247 (DUF247) proteins is involved in plant development and stress response. Rice is an important worldwide cereal crop, although an increasing number of DUF proteins have been identified, the understanding of DUF proteins is still very limited in rice.
RESULTS: In this study, we identified 69 genes that encode DUF247 proteins in the rice (Oryza sativa) genome by homology searches and domain prediction. All the OsDUF247 proteins were classified into four major groups (I, II, III and IV) by phylogenetic analysis. Remarkably, OsDUF247 genes clustered on the chromosomes solely show close phylogenetic relationships, suggesting that gene duplications have driven the expansion of the DUF247 gene family in the rice genome. Tissue profile analysis showed that most DUF247 genes expressed at constitutive levels in seedlings, roots, stems, and leaves, except for seven genes (LOC_Os01g21670, LOC_Os03g19700, LOC_Os05g04060, LOC_Os08g26820, LOC_Os08g26840, LOC_Os08g26850 and LOC_Os09g13410) in panicles. These seven genes were induced by various abiotic stress, including cold, drought, heat, hormone treatment, and especially salt, as demonstrated by further experimental analysis. DUF247 proteins contain transmembrane domains located on the membrane, suggesting their significant roles in rice development and adaptation to the environment.
CONCLUSIONS: These findings lay the foundation for functional characterizations of DUF247 genes to unravel their exact role in rice cultivars.

Kisspeptin (Kiss) and kisspeptin receptor (Kissr) system is a key regulator of GnRH expression in several vertebrates. The Indian catfish, Clarias magur, is popular in the Indian sub-continent, and a neo-type of the Asian catfish, C. batrachus. Catfish breeding is constrained as males do not release milt captivity with/without stimulation. Magur Kiss/Kissr system comprising of kiss1, kiss2, kissr1, and kissr2 genes was characterized for the first time. Full-length mRNA was sequenced using RACE PCR. Neighbor-joining tree of predicted proteins shows one clade of teleost orthologs. Magur whole genome (NCBI GenBank) has single copies of each gene, though yet unannotated/misannotated. Anomalies in the nomenclature of earlier sequences in GenBank were noted. Relative gene expression was profiled during various ontogenic stages, in six tissues including brain and gonads at maturity, and also in brains and gonads of premature and spent fish. Expression of gnrh1, gnrhr1, and gnrhr2 was estimated concomitantly. The kiss1 was the first to be twofold upregulated (P < 0.05) at 12 h post fertilization. Kiss/Kissr genes expressed primarily in the brain, ovary, and testis. Though kiss2 was 10 times higher than kiss1, only kiss1 showed significant modulation across stages and appears to be the active isotype that regulates GnRH in magur.

CONCLUSIONS: Two trans-isopentenyl diphosphate synthase and one squalene synthase genes were identified and proved to be involved in the triterpenoid biosynthesis in Platycodon grandiflorus. Platycodon grandiflorus is a commonly used traditional Chinese medicine. The main bioactive compounds of P. grandiflorus are triterpenoid saponins. The biosynthetic pathway of triterpenoid saponins in P. grandiflorus has been preliminarily explored. However, limited functional information on related genes has been reported. A total of three trans-isopentenyl diphosphate synthases (trans-IDSs) genes (PgFPPS, PgGGPPS1 and PgGGPPS2) and one squalene synthase (SQS) gene (PgSQS) in P. grandiflorus were screened and identified from transcriptome dataset. Subcellular localization of the proteins was defined based on the analysis of GFP-tagged. The activity of genes was verified in Escherichia coli, demonstrating that recombinant PgFPPS catalysed the production of farnesyl diphosphate. PgGGPPS1 produced geranylgeranyl diphosphate, whereas PgGGPPS2 did not exhibit catalytic activity. By structural identification of encoding genes, a transmembrane region was found at the C-terminus of the PgSQS gene, which produced an insoluble protein when expressed in E. coli but showed no apparent effect on the enzyme function. Furthermore, some triterpenoid saponin synthesis-related genes were discovered by combining the component content and the gene expression assays at the five growth stages of P. grandiflorus seedlings. The accumulation of active components in P. grandiflorus was closely associated with the expression level of genes related to the synthesis pathway.

BACKGROUND: The basidiomycete fungus, Ganoderma boninense is the main contributor to oil palm Basal Stem Rot (BSR) in Malaysia and Indonesia. Lanosterol 14α-Demethylase (ERG11) is a key enzyme involved in biosynthesis of ergosterol, which is an important component in the fungal cell membrane. The Azole group fungicides are effective against pathogenic fungi including G. boninense by inhibiting the ERG11 activity. However, the work on molecular characterization of G. boninense ERG11 is still unavailable today.
RESULTS: This study aimed to isolate and characterize the full-length cDNA encoding ERG11 from G. boninense. The G. boninense ERG11 gene expression during interaction with oil palm was also studied. A full-length 1860 bp cDNA encoding ERG11 was successfully isolated from G. boninense. The G. boninense ERG11 shared 91% similarity to ERG11 from other basidiomycete fungi. The protein structure homology modeling of GbERG11 was analyzed using the SWISS-MODEL workspace. Southern blot and genome data analyses showed that there is only a single copy of ERG11 gene in the G. boninense genome. Based on the in-vitro inoculation study, the ERG11 gene expression in G. boninense has shown almost 2-fold upregulation with the presence of oil palm.
CONCLUSIONS: This study provided molecular information and characterization study on the G. boninense ERG11 and this knowledge could be used to design effective control measures to tackle the BSR disease of oil palm.

UNASSIGNED: Newcastle disease (ND) is a viral infectious disease that affects commercial and native chickens, resulting in economic losses to the poultry industry. This study aimed to examine the viral strains circulating in commercial and native chickens by genetic characterization and observe the distribution of Newcastle disease virus (NDV) in chicken embryonic tissue.
UNASSIGNED: ND was detected using a quantitative reverse transcription-polymerase chain reaction. Genetic characterization of the fusion (F) and hemagglutinin-neuraminidase (HN) genes from the eight NDVs was performed using specific primers. The sequence was compared with that of other NDVs from GenBank and analyzed using the MEGA-X software. The distribution of NDV in chicken embryos was analyzed based on lesions and the immunopositivity in immunohistochemistry staining.
UNASSIGNED: Based on F gene characterization, velogenic NDV strains circulating in commercial and native chickens that showed varying clinical symptoms belonged to genotype VII.2. Lentogenic strains found in chickens without clinical symptoms were grouped into genotype II (unvaccinated native chickens) and genotype I (vaccinated commercial chickens). Amino acid variations in the HN gene, namely, the neutralization epitope and antigenic sites at positions 263 and 494, respectively, occurred in lentogenic strains. The NDV reaches the digestive and respiratory organs, but in lentogenic NDV does not cause significant damage, and hence embryo death does not occur.
UNASSIGNED: This study showed that velogenic and lentogenic NDV strains circulated in both commercial and native chickens with varying genotypes. The virus was distributed in almost all organs, especially digestive and respiratory. Organ damage in lentogenic infection is not as severe as in velogenic NDV. Further research is needed to observe the distribution of NDV with varying pathogenicity in chickens.

Insects have been recognized as sustainable alternative sources of nutrients for food and feed. The Black Soldier Fly (BSF), Hermetia illucens, is a particularly promising species for its great potential in the waste valorization to produce, during the bioconversion process, high-value fat and proteins that currently represent a valuable source for fish feed. The present study aims to evaluate the efficiency to use substrate proteins in two different BSF developmental stages as sustainable biotechnological tools for vegetable waste management. We provide insights into the nutritional values of both V instar larvae and prepupae in terms of valuable amino acids with special focus on taurine, a crucial nutrient for fish. Moreover, we cloned four key genes from BSF involved in the taurine biosynthesis pathway, 2-aminoethanethiol dioxygenase (Hiado), cysteine dioxygenase (Hicdo), cysteine sulfonate decarboxylase (Hicsad), and glutamate decarboxylase (Higad). The gene expression analysis in larvae and prepupae by qPCR showed development-specific profiles suggesting they influence the taurine content during BSF development. These findings showed peculiar phenotypes in larvae and prepupae that can be selected for different biotechnological applications as sustainable source of relevant amino acids and taurine to support the increasing demand for animal feed and aquafeed in the next decades.

The rust pathogens are one of the most complex fungi in the Basidiomycetes. The development of genomic resources for rust and other plant pathogens has opened the opportunities for functional genomics of fungal genes. Despite significant progress in the field of fungal genomics, functional characterization of the genome components has lacked, especially for the rust pathogens. Their obligate nature and lack of standard stable transformation protocol are the primary reasons for rusts to be one of the least explored genera despite its significance. In the recently sequenced rust genomes, a vast catalogue of predicted effectors and pathogenicity genes have been reported. However, most of these candidate genes remained unexplored due to the lack of suitable characterization methods. The heterologous expression of putative effectors in Nicotiana benthamiana and Arabidopsis thaliana has proved to be a rapid screening method for identifying the role of these effectors in virulence. However, no fungal system has been used for the functional validation of these candidate genes. The smuts, from the evolutionary point of view, are closely related to the rust pathogens. Moreover, they have been widely studied and hence could be a suitable model system for expressing rust fungal genes heterologously. The genetic manipulation methods for smuts are also well standardized. Complementation assays can be used for functional validation of the homologous genes present in rust and smut fungal pathogens, while the species-specific proteins can be expressed in the mutant strains of smut pathogens having reduced or no virulence for virulence analysis. We propose that smuts, especially Ustilago maydis, may prove to be a good model system to characterize rust effector proteins in the absence of methods to manipulate the rust genomes directly.

Recent advances in genomics present new opportunities for enhancing knowledge about gene regulation and function across a wide spectrum of organisms and species. Understanding and evaluating this information at the individual gene level is challenging, and not only requires extracting, collating and interpreting data from public genetic repositories, but also recognizing that much of the information has been developed through implementation of computationally based exon-calling algorithms, and thus may be inaccurate. Moreover, as these data usually have not been validated experimentally, results also may be incomplete and incorrect. This has created a quality-control problem for scientists who want to use individual gene-specific information in their research. Here, I describe a simple experimental strategy that takes advantage of the large amounts of untapped primary experimental data for characterizing gene expression that have been deposited in the Sequence Read Archive of the National Center for Biotechnology Information. The approach consists of a readily adaptable pipeline that may be used to confirm exons, to define 5\' and 3\' un-translated regions and the beginnings and ends of individual genes, and to quantify alternative RNA splicing. The series of experimental strategies described offers effective replacements for older molecular biological methods, and can rapidly and reproducibly resolve major gene mapping problems.