Transcriptome dataset of Metroxylon sagu palms from multiple sago plantations in Sarawak.

OBJECTIVE: Sago palm (Metroxylon sagu Rottb.) is one of the most important economic crops abundantly found in Mukah, Sarawak, Malaysia. The robustness of the palm triggered the Sarawak government's selection as one of the state's commodity crops, with the opening of several sago palm plantations. However, stunted (non-trunking) palms were reported in several sago palm plantations despite attaining a maturity period of more than ten years after cultivation. Research targeting this problem has been conducted in various fields, yet information on molecular mechanisms is still scarce. This study aimed to determine the genes responsible for sago palm's normal phenotype (trunking) by attaining leaf transcriptomes from samples of all trunking sago palms from different sago palm plantations. DATA DESCRIPTION: The conventional CTAB method was employed in the present investigation to extract total RNA from leaf tissues. Transcriptome sequencing was conducted on the Illumina NovaSeq 6000 platform. Differential expression analysis was performed using the DESeq2 package. A total of 6,119 differentially expressed genes, comprising 4,384 downregulated and 1,735 upregulated genes, were expressed in all three sago palm datasets. The datasets provide insights into the commonly expressed genes among trunking sago palms.

Transcriptome dataset of sago palm in peat soil.

Sago palm (Metroxylon sagu Rottb.) is an important agricultural starch-producing palm that contributes to Malaysia's economics, especially in the State of Sarawak, Malaysian Borneo. In this palm tree, the central part of the plant storage-starch. Under normal condition, sago palm develop its trunk after 4-5 years being planted. However, sago palms planted on deep-peat soil failed to develop their trunk even after 17 years of being planted. This phenomenon is known as 'non-trunking', which eliminates the economic value of the palms. Numerous research has been done to address the phenomenon, but the molecular mechanisms of sago palm responding toward the responsible stresses are still lacking. Therefore, in this study, leaf samples were collected from trunking (normal) and non-trunking sago palms planted on peat soil for total RNA extraction, followed by next-generation sequencing using the BGISEQ-500 platform. The raw reads were cleaned, and de novo assembled using TRINITY software package. A total of 40.11 Gb bases were sequenced from the sago palm leaf samples. The assembled sequence produced 102,447 unigenes, with N50 score 1809 bp and GC ratio of 44.34%. The alignment of unigenes with seven functional databases (NR, NT, GO, KOG, KEGG, SwissProt and InterPro) resulted in the annotation of 65,523 (63.96%) unigenes. Functional annotation results in the detection of 46,335 coding DNA sequences by Transdecoder. A total of 30,039 simple-sequence repeats distributed on 21,676 unigenes were detected using Primer3 software, and 2355 transcription factor coding unigenes were predicted using getorf and hmmseach software. This work is registered under NCBI BioProject PRJNA781491. The raw RNA sequencing data are available in Sequence Read Archive (SRA) database with accession numbers SRX13165895, SRX13165896, SRX13165897, SRX13165898, SRX13165899, and SRX13165900. Gene expression and annotation information are accessible in public functional genomics data repository Gene Expression Omnibus (GEO) with accession number GSE189085.

First high-quality genome assembly data of sago palm (Metroxylon sagu Rottboll).

The sago palm (Metroxylon sagu Rottboll) is a tropical halophytic starch-producing, economically important crop palm mainly located in Southeast Asian countries. Recently, a genome survey was conducted on this palm using the Illumina sequencing platform, with a very low (21.5%) BUSCO genome completeness score, and most of them (∼78%) are either fragmented or missing. Thus, in this study, the sago palm genome completeness was further improved with the utilization of the Nanopore sequencing platform that produced longer reads. A hybrid genome assembly was conducted, and the outcome was a much complete sago palm genome with BUSCO completeness achieved at as high as 97.9%, with only ∼2% of them either fragmented or missing. The estimated genome size of the sago palm is 509,812,790 bp in this study. A sum of 33,242 protein-coding genes was revealed from the sago palm genome and around 96.39% of them had been functionally annotated. An investigation on the carbohydrate metabolism KEGG pathways also unearthed that starch synthesis was one of the major sago palm activities. The genome data obtained from this work is indispensable for future molecular evolutionary and genome-wide association studies on the economically important sago palm.

Proteomics of Sago Palm Towards Identifying Contributory Proteins in Stress-Tolerant Cultivar.

Metroxylon sagu Rottb. or locally known as sago palm is a tropical starch crop grown for starch production in commercial plantations in Malaysia, especially in Sarawak, East Malaysia. This plant species accumulate the highest amount of edible starch compared to other starch-producing crops. However, the non-trunking phenomenon has been observed to be one of the major issues restricting the yield of sago palm starch. In this study, proteomics approach was utilised to discover differences between trunking and non-trunking proteomes in sago palm leaf tissues. Total protein from 16 years old trunking and non-trunking sago palm leaves from deep peat area were extracted with PEG fractionation extraction method and subjected to two-dimensional gel electrophoresis (2D PAGE). Differential protein spots were subjected to MALDI-ToF/ToF MS/MS. Proteomic analysis has identified 34 differentially expressed proteins between trunking and non-trunking sago samples. From these protein spots, all 19 proteins representing different enzymes and proteins have significantly increased in abundance in non-trunking sago plant when subjected to mass spectrometry. The identified proteins mostly function in metabolic pathways including photosynthesis, tricarboxylic acid cycle, glycolysis, carbon utilization and oxidative stress. The current study indicated that the several proteins identified through differentially expressed proteome contributed to physical differences in trunking and non-trunking sago palm.

Characterization and expression in Pichia pastoris of a raw starch degrading glucoamylase (GA2) derived from Aspergillus flavus NSH9.

The Aspergillus flavus NSH9 gene, encoding a pH and thermostable glucoamylase with a starch binding domain (SBD), was expressed in Pichia pastoris to produce recombinant glucoamylase (rGA2). The full-length glucoamylase gene (2039 bp), and cDNA (1839 bp) encode a 612 amino acid protein most similar to glucoamylase from Aspergillus oryzae RIB40; the first 19 amino acids are presumed to be a signal peptide for secretion, and the SBD is at the C-terminal. The cDNA was successfully secreted by Pichia at 8.23 U mL-1, and the rGA2 was found to be: a 80 kDa monomer, stable from pH 3.0-9.0, with optimum catalytic activity at pH 5.0, active at temperatures up to 80°C (rGA2 retained 58% of its activity after 60 min of incubation at 70°C), and metal ions such as Na+, K+, Ca++ and Mg++ enhanced rGA2 enzyme activity. The starch degrading ability of rGA2 was also observed on raw sago starch and where prolonged incubation generated larger, deeper, holes on the starch granules, indicating rGA2 is an excellent candidate for industrial starch processing applications.

Celiac disease: What the Indian pediatricians know about the disease.

To ascertain the knowledge, awareness, and practices pertaining to celiac disease (CD) among the Indian pediatricians. A survey link containing a questionnaire was shared through electronic mail using a pediatric database. The survey was kept active for 6 months; all responses received at the end of the survey were analyzed. Two hundred and seventy one pediatricians out of more than 10,000 chose to respond to the survey. Most pediatricians agreed that more patients with CD are being diagnosed than earlier. The reasons for higher detection of CD were perceived to be higher index of clinical suspicion by pediatricians (86.7%) followed by increased awareness among parents (45.8%). Most pediatricians opined that clinical manifestations which prompted to a diagnosis of CD were failure to thrive (96.2%) and chronic diarrhea (81.4%). Knowledge about atypical manifestations of celiac disease was low.  Though knowledge about the common association of CD with type 1 diabetes (62.1%) and autoimmune hepatitis (55.8%) was there, awareness about its association with other uncommon conditions was lacking. Though 68% of the pediatricians were of the opinion that the confirmation of diagnosis by a mucosal biopsy is necessary, 26.5% of respondents believed that only a positive serology was sufficient for a diagnosis. A trial of gluten-free diet (GFD) was thought to be a logical step if serology was positive by 31.3% of respondents. While 87.7% of pediatricians advocated lifelong adherence to GFD, 12.3% felt that GFD could be discontinued in the future. This web-based survey revealed that though pediatricians are seeing increasing number of celiac disease patients, there is a need to increase awareness regarding the disease, its associated conditions, the need for mucosal biopsy to confirm the diagnosis and the necessity of lifelong adherence to GFD.

Purification of an alpha amylase from Aspergillus flavus NSH9 and molecular characterization of its nucleotide gene sequence.

In this study, an alpha-amylase enzyme from a locally isolated Aspergillus flavus NSH9 was purified and characterized. The extracellular α-amylase was purified by ammonium sulfate precipitation and anion-exchange chromatography at a final yield of 2.55-fold and recovery of 11.73%. The molecular mass of the purified α-amylase was estimated to be 54 kDa using SDS-PAGE and the enzyme exhibited optimal catalytic activity at pH 5.0 and temperature of 50 °C. The enzyme was also thermally stable at 50 °C, with 87% residual activity after 60 min. As a metalloenzymes containing calcium, the purified α-amylase showed significantly increased enzyme activity in the presence of Ca2+ ions. Further gene isolation and characterization shows that the α-amylase gene of A. flavus NSH9 contained eight introns and an open reading frame that encodes for 499 amino acids with the first 21 amino acids presumed to be a signal peptide. Analysis of the deduced peptide sequence showed the presence of three conserved catalytic residues of α-amylase, two Ca2+-binding sites, seven conserved peptide sequences, and several other properties that indicates the protein belongs to glycosyl hydrolase family 13 capable of acting on α-1,4-bonds only. Based on sequence similarity, the deduced peptide sequence of A. flavus NSH9 α-amylase was also found to carry two potential surface/secondary-binding site (SBS) residues (Trp 237 and Tyr 409) that might be playing crucial roles in both the enzyme activity and also the binding of starch granules.

Combined Overlap Extension PCR Method for Improved Site Directed Mutagenesis.

The combined overlap extension PCR (COE-PCR) method developed in this work combines the strengths of the overlap extension PCR (OE-PCR) method with the speed and ease of the asymmetrical overlap extension (AOE-PCR) method. This combined method allows up to 6 base pairs to be mutated at a time and requires a total of 40-45 PCR cycles. A total of eight mutagenesis experiments were successfully carried out, with each experiment mutating between two to six base pairs. Up to four adjacent codons were changed in a single experiment. This method is especially useful for codon optimization, where doublet or triplet rare codons can be changed using a single mutagenic primer set, in a single experiment.

Application of Response Surface Methodology for Optimizing Process Parameters in the Production of Amylase by Aspergillus flavusNSH9 under Solid State Fermentation

Amylase is recognized as one of the important commercial enzymes. This group of enzymes has the ability in hydrolyzing starch into smaller oligosacharides. The present work aimed to determine the optimum fermentation conditions for maximum production of crude amylase enzyme by Aspergillus flavus NSH9 employing response surface methodology (RSM).Central composite design (CCD) was applied to determine the optimal fermentation condition with respect to the four main process parameters such as temperature, initial moisture content, pH and the incubation period. Solid state fermentation (SSF) was performed using 5.0 g of sago hampas inoculated with 1x107sporesmL-1following the experimental design obtained using CCD and further optimized by RSM. The initial moisture, pH and temperature showed significant effect on the amylase production (p<0.05). The maximum amylase activity produced was achieved and recorded as 1.055 ± 0.03U mL-1after four days of fermentation period with 100% (v/v) moisture holding capacity, pH 6.5 and temperature at 28°C. The optimum fermentation conditions for amylase production was determined with A. flavusNSH9 on sago hampas.

N,N'-Bis(phenyl-carbamo-thio-yl)benzene-1,3-dicarboxamide.

The asymmetric unit of the title compound, C22H18N4O2S2, contains two mol-ecules. In one of them, the dihedral angles between the central benzene ring and the phenyl rings are 16.97 (8) and 20.97 (8)°, while the phenyl rings make a dihedral angle of 37.87 (8)°. In the other mol-ecule, the corresponding values are 34.92 (7), 53.90 (7) and 60.68 (8)°, respectively. In each mol-ecule, two intra-molecular N-H⋯O hydrogen bonds generate S(6) rings and a short C-H⋯S contact also occurs. In the crystal, N-H⋯S, N-H⋯O, C-H⋯O and C-H⋯S inter-actions link the mol-ecules into a three-dimensional network.

(E)-1-[4-(Hex-yloxy)phen-yl]-3-(2-hy-droxy-phen-yl)prop-2-en-1-one.

In the title compound, C(21)H(24)O(3), the enone moiety adopts an s-cis conformation and the dihedral angle between the benzene rings is 12.89 (6)°. The hex-yloxy tail adopts an extended conformation. In the crystal, inversion dimers are linked by pairs of O-H⋯O hydrogen bonds and pairs of C-H⋯O inter-actions, forming two R(2) (2)(7) and one R(2) (2)(10) loops. The dimers are then arranged into sheets lying parallel to (201) and weak C-H⋯π inter-actions consolidate the packing.

(E)-1-(3-Hy-droxy-phen-yl)-3-[4-(tetra-dec-yl-oxy)phen-yl]prop-2-en-1-one.

In the title compound, C(29)H(40)O(3), the enone moiety adopts an s-cis conformation. The dihedral angle between the benzene rings is 4.33 (5)° The least-squares mean line through the tetra-decyl side chain forms a dihedral angle of 83.99 (7)° with the normal to the attached benzene ring. In the crystal, O-H⋯O and C-H⋯O hydrogen bonds involving the keto and the hy-droxy O atoms form ribbons along [-41-1]. The crystal structure also features C-H⋯π inter-actions.

(E)-1-(4-Decyl-oxyphen-yl)-3-(2-hy-droxy-phen-yl)prop-2-en-1-one.

In the title compound, C(25)H(32)O(3), the enone group adopts an s-cis conformation. The alk-oxy chain is in an all-trans conformation. The dihedral angle between the benzene rings is 7.86 (5)°. In the crystal, mol-ecules are connected by pairs of O-H⋯O hydrogen bonds, forming inversion dimers and giving R(2) (2)(10) rings. Within these dimers, weak C-H⋯O hydrogen bonds form two R(2) (2)(7) rings. In the crystal, the approximately planar mol-ecules [largest deviation for an atom being 0.4737 (12) Šfor the terminal C atom of the alk-oxy chain] are arranged in sheets parallel to (20-1). Weak C-H⋯π inter-actions are also observed.

Synthesis and antimicrobial studies of hydroxylated chalcone derivatives with variable chain length.

A series of (E)-1-(4-alkyloxyphenyl)-3-(hydroxyphenyl)-prop-2-en-1-one have been successfully synthesised via Claisen-Schmidt condensation. The synthesised chalcone derivatives consisted of hydroxyl groups at either ortho, meta or para position and differed in the length of the alkyl groups, C (n) H(2) (n) (+1,) where n = 6, 10, 12 and 14. The structures of all compounds were defined by elemental analysis, IR, (1)H- and (13)C-NMR. The antimicrobial studies were carried out against wild-type Escherichia coli American Type Culture Collection 8739 to evaluate the effect of the hydroxyl and the alkyl groups of the synthesised chalcones. All the synthesised compounds have shown significant antimicrobial activities. The optimum inhibition was dependent on the position of the hydroxyl group as well as the length of the alkyl chains.

(E)-1-[4-(Hex-yloxy)phen-yl]-3-(3-hy-droxy-phen-yl)prop-2-en-1-one.

There are two mol-ecules in the asymmetric unit of the title compound, C(21)H(24)O(3), in which the dihedral angles between the aromatic rings are 6.4 (1) and 7.0 (1)°. The enone moiety of both mol-ecules adopts an s-cis configuration. In the crystal, inter-molecular O-H⋯O and C-H⋯O inter-actions to the same acceptor O atom generate R(2) (1)(6) ring motifs and further C-H⋯O inter-actions generate R(2) (2)(8) ring motifs. Topologically, the R(2) (1)(6) and R(2) (2)(8) ring motifs are arranged alternately, forming [001] chains of mol-ecules. The crystal structure is further stabilized by C-H⋯π inter-actions.

(E)-1-[4-(Hex-yloxy)phen-yl]-3-(4-hy-droxy-phen-yl)prop-2-en-1-one.

In the title compound, C(21)H(24)O(3), inter-molecular O-H⋯O and C-H⋯O inter-actions form bifurcated acceptor bonds, generating R(2) (1)(6) ring motifs. These ring motifs link the mol-ecules into extended chains along [010]. The crystal structure is further stabilized by C-H⋯π inter-actions.

(E)-3-(4-Hexyl-oxyphen-yl)-1-(3-hydroxy-phen-yl)prop-2-en-one.

In the title compound, C(21)H(24)O(3), the enone unit is in the s-cis configuration. The dihedral angle between the benzene rings is 2.18 (4)°. In the crystal, mol-ecules are linked by pairs of O-H⋯O inter-molecular hydrogen bonds, forming inversion dimers. The crystal structure is also consolidated by C-H⋯π inter-actions.

(E)-1-(4-Decyl-oxyphen-yl)-3-(4-hydroxy-phen-yl)prop-2-en-1-one.

In the title compound, C(25)H(32)O(3), the asymmetric unit contains two crystallographically independent mol-ecules: both enone groups adopt an s-cis configuration. In the crystal, O-H⋯O and C-H⋯O inter-molecular inter-actions form bifurcated hydrogen bonds, which generate R(1) (2)(6) ring motifs. These inter-molecular inter-actions link the mol-ecules into one-dimensional chains along the [10] direction. The crystal structure is further stabilized by C-H⋯π inter-actions.

(E)-3-(4-Decyl-oxyphen-yl)-1-(2-hydroxy-phen-yl)prop-2-en-1-one.

In the title compound, C(25)H(32)O(3), the enone group is in an s-cis configuration. The dihedral angle between the benzene rings is 8.84 (7)°. An intra-molecular O-H⋯O inter-action between the keto and hydr-oxy groups forms an S(6) ring motif. Inter-molecular C-H⋯O inter-actions link the mol-ecules into supra-molecular chains along the c axis which are subsequently stacked down the b axis; the crystal structure is further consolidated by C-H⋯π inter-actions.

(E)-3-[4-(Hex-yloxy)phen-yl]-1-(2-hydroxy-phen-yl)prop-2-en-1-one.

In the title compound, C(21)H(24)O(3), the conformation of the enone group is s-cis. The benzene rings are inclined at an angle of 7.9 (1)°. The alk-oxy tail is planar, with a maximum deviation from the least-squares plane of 0.009 (2) Å, and adopts a trans conformation throughout. An intra-molecular O-H⋯O inter-action between the keto and hydr-oxy groups forms S(6) ring motifs. In the crystal, mol-ecules are arranged in a head-to-tail manner down the a axis and are subsequently stacked along the b axis, forming mol-ecular sheets parallel to the ab plane. The crystal structure is further stabilized by weak C-H⋯π inter-actions and short C⋯O [3.376 (2) Å] contacts.