Chromosomal Location Research Articles

CRISPR-COPIES: an in silico platform for discovery of neutral integration sites for CRISPR/Cas-facilitated gene integration.

The CRISPR/Cas system has emerged as a powerful tool for genome editing in metabolic engineering and human gene therapy. However, locating the optimal site on the chromosome to integrate heterologous genes using the CRISPR/Cas system remains an open question. Selecting a suitable site for gene integration involves considering multiple complex criteria, including factors related to CRISPR/Cas-mediated integration, genetic stability, and gene expression. Consequently, identifying such sites on specific or different chromosomal locations typically requires extensive characterization efforts. To address these challenges, we have developed CRISPR-COPIES, a COmputational Pipeline for the Identification of CRISPR/Cas-facilitated intEgration Sites. This tool leverages ScaNN, a state-of-the-art model on the embedding-based nearest neighbor search for fast and accurate off-target search, and can identify genome-wide intergenic sites for most bacterial and fungal genomes within minutes. As a proof of concept, we utilized CRISPR-COPIES to characterize neutral integration sites in three diverse species: Saccharomyces cerevisiae, Cupriavidus necator, and HEK293T cells. In addition, we developed a user-friendly web interface for CRISPR-COPIES (https://biofoundry.web.illinois.edu/copies/). We anticipate that CRISPR-COPIES will serve as a valuable tool for targeted DNA integration and aid in the characterization of synthetic biology toolkits, enable rapid strain construction to produce valuable biochemicals, and support human gene and cell therapy applications.

Genome-wide identification and expression analysis of autophagy-related genes (ATG) in Gossypium spp. reveals their crucial role in stress tolerance

Autophagy is an evolutionarily conserved process mediated by AuTophaGy (ATG) genes, in which cellular components are degraded and recycled within vacuoles or lysosome. Yeast ATG was extensively studied; their homologs are reported in plants including Arabidopsis. However, they have not been reported in cotton. In the present study, the ATG cascade genes were studied in four Gossypium species, namely G. hirsutum, G. arboreum, G. herbaceum, and G. raimondii. A total of 40 GhirATG, 33 GaATG, 34 GherbATG, and 25 GrATG genes were identified in these species. These ATG sequences exhibited conserved ATG domains and other overlapping domains, including Ras, Pkinase, Snf7, WD40, Glyco_hydro_17, and bZIP_1, and were named based on their phylogenetic relationships with Arabidopsis ATGs. A phylogenetic analysis revealed evolutionary relationships among ATG genes among these species, as well as significant sub-clades indicating patterns of gene retention. Based on synonymous and non-synonymous substitutions, cotton groups diverged at different times from Arabidopsis. Gene structure analysis and chromosome localization showed exon-intron patterns and locations. Expression analysis of ATGs in different tissues during development and under stress conditions indicated their functional specificity. The ATG8 subclass genes were significantly expressed under abiotic stress conditions, indicating their role in stress response. In addition, qRT-PCR analysis of selected 12 GhirATGs in leaves treated with stress elicitors, MeJA, SA, NaCl, and PEG for varying time intervals showed a pattern of their significant upregulation. The findings contribute to our understanding of autophagy-related processes in cotton, including their potential role in stress response and development.

Quantum Evolution and Genetic Mutations

One of the important problems in quantum biology has been explained in this work. It is the main reason for genetic mutation that is known as the origin of evolution in the organisms. An approach to clarify the main reason of mutation is quantum mechanics. A brief history of genetics, new-Darwinism and Mendelian inheritance from ancient time to the twentieth century has been presented. Meaning and location of chromosomes and genes in cells, DNA structure, the replication mechanism, genetic code, tautomeric form and application of quantum mechanics have been described. Finally, we present the importance of quantum tunnelling to play a main role in the mutation.

In Silico Identification and Characterization of Rare Cold Inducible 2 (RCI2) Gene Family in Cotton.

RCI2/PMP3s are involved in biotic and abiotic stresses and have an influence on the regulation of many genes. RCI2/PMP3 genes, which particularly encode small membrane proteins of the PMP3 family, are involved in abiotic stress responses in plants. In this work, in silico studies were used to investigate RCI2's potential function in stress tolerance and organogenesis. We conducted an extensive study of the RCI2 gene family and revealed 36 RCI2 genes from cotton species that were distributed over 36 chromosomes of the cotton genome. Functional and phylogenetic examination of the RCI2/PMP3 gene family has been studied in Arabidopsis, but in cotton, the RCI2/PMP3 genes have not yet been studied. Phylogenetic and sequencing studies revealed that cotton RCI2s are conserved, with most of them categorized into six distinct clades. A chromosome distribution and localization study indicated that cotton RCI2 genes were distributed unevenly on 36 chromosomes with segmental duplications, suggesting that the cotton RCI2 family is evolutionarily conserved. Many cis-elements related to stress responsiveness, development, and hormone responsiveness were detected in the promoter regions of the cotton RCI2. Moreover, the 36 cotton RCI2s revealed tissue-specific expression patterns in the development of cotton performed by transcriptome analysis. Gene structure analysis indicated that nearly all RCI2 genes have two exons and one intron. All of the cotton RCI2 genes were highly sensitive to drought, abscisic acid, salt, and cold treatments, demonstrating that they may be employed as genetic objects to produce stress-resistant plants.

Theoretical Analysis and Expression Profiling of 17β-Hydroxysteroid Dehydrogenase Genes in Gonadal Development and Steroidogenesis of Leopard Coral Grouper (Plectropomus leopardus).

The differentiation and developmental trajectory of fish gonads, significantly important for fish breeding, culture, and production, has long been a focal point in the fields of fish genetics and developmental biology. However, the mechanism of gonadal differentiation in leopard coral grouper (Plectropomus leopardus) remains unclear. This study investigates the 17β-Hydroxysteroid Dehydrogenase (Hsd17b) gene family in P. leopardus, with a focus on gene characterization, expression profiling, and functional analysis. The results reveal that the P. leopardus's Hsd17b gene family comprises 11 members, all belonging to the SDR superfamily. The amino acid similarity is only 12.96%, but conserved motifs, such as TGxxxGxG and S-Y-K, are present in these genes. Hsd17b12a and Hsd17b12b are unique homologs in fish, and chromosomal localization has confirmed that they are not derived from different transcripts of the same gene, but rather are two independent genes. The Hsd17b family genes, predominantly expressed in the liver, heart, gills, kidneys, and gonads, are involved in synthesizing or metabolizing sex steroid hormones and neurotransmitters, with their expression patterns during gonadal development categorized into three distinct categories. Notably, Hsd17b4 and Hsd17b12a were highly expressed in the testis and ovary, respectively, suggesting their involvement in the development of reproductive cells in these organs. Fluorescence in situ hybridization (FISH) further indicated specific expression sites for these genes, with Hsd17b4 primarily expressed in germ stem cells and Hsd17b12a in oocytes. This comprehensive study provides foundational insights into the role of the Hsd17b gene family in gonadal development and steroidogenesis in P. leopardus, contributing to the broader understanding of fish reproductive biology and aquaculture breeding.

GhBRX.1, GhBRX.2, and GhBRX4.3 improve resistance to salt and cold stress in upland cotton.

Abiotic stress during growth readily reduces cotton crop yield. The different survival tactics of plants include the activation of numerous stress response genes, such as BREVIS RADIX (BRX). In this study, the BRX gene family of upland cotton was identified and analyzed by bioinformatics method, three salt-tolerant and cold-resistant GhBRX genes were screened. The expression of GhBRX.1, GhBRX.2 and GhBRXL4.3 in upland cotton was silenced by virus-induced gene silencing (VIGS) technique. The physiological and biochemical indexes of plants and the expression of related stress-response genes were detected before and after gene silencing. The effects of GhBRX.1, GhBRX.2 and GhBRXL4.3 on salt and cold resistance of upland cotton were further verified. We discovered 12, 6, and 6 BRX genes in Gossypium hirsutum, Gossypium raimondii and Gossypium arboreum, respectively. Chromosomal localization indicated that the retention and loss of GhBRX genes on homologous chromosomes did not have a clear preference for the subgenomes. Collinearity analysis suggested that segmental duplications were the main force for BRX gene amplification. The upland cotton genes GhBRX.1, GhBRX.2 and GhBRXL4.3 are highly expressed in roots, and GhBRXL4.3 is also strongly expressed in the pistil. Transcriptome data and qRT‒PCR validation showed that abiotic stress strongly induced GhBRX.1, GhBRX.2 and GhBRXL4.3. Under salt stress and low-temperature stress conditions, the activities of superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT) and the content of soluble sugar and chlorophyll decreased in GhBRX.1-, GhBRX.2- and GhBRXL4.3-silenced cotton plants compared with those in the control (TRV: 00). Moreover, GhBRX.1-, GhBRX.2- and GhBRXL4.3-silenced cotton plants exhibited greater malondialdehyde (MDA) levels than did the control plants. Moreover, the expression of stress marker genes (GhSOS1, GhSOS2, GhNHX1, GhCIPK6, GhBIN2, GhSnRK2.6, GhHDT4D, GhCBF1 and GhPP2C) decreased significantly in the three target genes of silenced plants following exposure to stress. These results imply that the GhBRX.1, GhBRX.2 and GhBRXL4.3 genes may be regulators of salt stress and low-temperature stress responses in upland cotton.

A Breast Cancer Candidate Locus at 6q Narrowed to 6q15-q21.

Although a number of high-risk breast cancer genes have been identified, including BRCA1 and BRCA2, the risk profile of many high-risk families cannot be explained using known breast cancer genes. Previously, we have shown strong indications of new breast cancer risk loci at chromosomes 2p, 6q, and 14q in a family of six generations including 10 breast cancer cases. In this study, we identified and traced four new family branches descending from siblings of the parents in the top generation of the studied family. One distantly related branch included four breast cancer cases, two of whom were diagnosed at age < 45 years. DNA samples from the cases were typed at selected polymorphic markers from all three chromosome loci, to test identical origin of the haplotypes. All four cases were shown to segregate a common 6q haplotype with a region identical to the previously identified 6q haplotype. The data strongly support a new breast cancer locus at 6q, and narrow it down to a 17 MB interval at 6q15-q21.

Genetic characterization of multidrug-resistant Escherichia coli harboring colistin-resistant gene isolated from food animals in food supply chain.

Colistin is widely used for the prophylaxis and treatment of infectious disease in humans and livestock. However, the global food chain may actively promote the dissemination of colistin-resistant bacteria in the world. Mobile colistin-resistant (mcr) genes have spread globally, in both communities and hospitals. This study sought to genomically characterize mcr-mediated colistin resistance in 16 Escherichia coli strains isolated from retail meat samples using whole genome sequencing with short-read and long-read platforms. To assess colistin resistance and the transferability of mcr genes, antimicrobial susceptibility testing and conjugation experiments were conducted. Among the 16 isolates, 11 contained mcr-1, whereas three carried mcr-3 and two contained mcr-1 and mcr-3. All isolates had minimum inhibitory concentration (MIC) for colistin in the range 1-64 μg/mL. Notably, 15 out of the 16 isolates demonstrated successful transfer of mcr genes via conjugation, indicative of their presence on plasmids. In contrast, the KK3 strain did not exhibit such transferability. Replicon types of mcr-1-containing plasmids included IncI2 and IncX4, while IncFIB, IncFII, and IncP1 contained mcr-3. Another single strain carried mcr-1.1 on IncX4 and mcr-3.5 on IncP1. Notably, one isolate contained mcr-1.1 located on a chromosome and carrying mcr-3.1 on the IncFIB plasmid. The chromosomal location of the mcr gene may ensure a steady spread of resistance in the absence of selective pressure. Retail meat products may act as critical reservoirs of plasmid-mediated colistin resistance that has been transmitted to humans.

Identification and expression of genome of uridine diphosphate glycosyltransferase (UGT) gene family from Chrysanthemum indicum

Uridine diphosphate glycosyltransferase(UGT) is involved in the glycosylation of a variety of secondary metabolites in plants and plays an important role in plant growth and development and regulation of secondary metabolism. Based on the genome of a diploid Chrysanthemum indicum, the UGT gene family from Ch. indicum was identified by bioinformatics methods, and the physical and chemical properties, subcellular localization prediction, conserved motif, phylogeny, chromosome location, gene structure, and gene replication events of UGT protein were analyzed. Transcriptome and real-time fluorescence quantitative polymerase chain reaction(PCR) were used to analyze the expression pattern of the UGT gene in flowers and leaves of Ch. indicum. Quasi-targeted metabolomics was used to analyze the differential metabolites in flowers and leaves. The results showed that a total of 279 UGT genes were identified in the Ch. indicum genome. Phylogenetic analysis showed that these UGT genes were divided into 8 subfamilies. Members of the same subfamily were distributed in clusters on the chromosomes. Tandem duplications were the main driver of the expansion of the UGT gene family from Ch. indicum. Structural domain analysis showed that 262 UGT genes had complete plant secondary metabolism signal sequences(PSPG box). The analysis of cis-acting elements indicated that light-responsive elements were the most ubiquitous elements in the promoter regions of UGT gene family members. Quasi-targeted metabolome analysis of floral and leaf tissue revealed that most of the flavonoid metabolites, including luteolin-7-O-glucoside and kaempferol-7-O-glucoside, had higher accumulation in flowers. Comparative transcriptome analysis of flower and leaf tissue showed that there were 72 differentially expressed UGT genes, of which 29 genes were up-regulated in flowers, and 43 genes were up-regulated in leaves. Correlation network and phylogenetic analysis showed that CindChr9G00614970.1, CindChr2G00092510.1, and CindChr2G00092490.1 may be involved in the synthesis of 7-O-flavonoid glycosides in Ch. indicum, and real-time fluorescence quantitative PCR analysis further confirmed the reliability of transcriptome data. The results of this study are helpful to understand the function of the UGT gene family from Ch. indicum and provide data reference and theoretical basis for further study on the molecular regulation mechanism of flavonoid glycosides synthesis in Ch. indicum.

Development and evaluation of a novel single nucleotide polymorphism panel for North American bison.

Genome-wide single nucleotide polymorphism (SNP) genotyping platforms have become increasingly popular in characterizing livestock and wildlife populations, replacing traditional methods such as microsatellite fragment analysis. Herein, we report the development and evaluation of a novel bison SNP panel for population management and conservation. Initially, 2474 autosomal SNPs were selected from existing bison whole-genome sequences and variable sites among bison on the GGSP bovine 50K Chip, based on minor allele frequency, data completeness, and chromosome location. Additionally, 20 mitochondrial SNPs were chosen to identify known mitochondrial haplotypes in bison according to previous research. The SNPs were further evaluated using genotyping-by-sequencing with 190 bison, representing the historical lineages that survived the major population crash of the late 1800s. Variants with high potential for genotyping error were filtered out, and the remaining SNPs were placed on a custom Illumina™ array. The final panel consisting of 798 autosomal and 13 mitochondrial SNPs was used to establish baseline genetic parameters, compare populations, and assign mitochondrial haplotypes in 995 bison across ten populations. These SNPs were also found to be highly informative for individual animal identification and parentage assignment. This SNP panel provides a powerful new method to establish a baseline for estimating genetic health of bison populations and a new tool for bison managers to make informed management decisions based on genetic information specific to their populations.

Genome-Wide Identification of Expansins in Rubus chingii and Profiling Analysis during Fruit Ripening and Softening

Improving fruit size or weight, firmness, and shelf life is a major target for horticultural crop breeding. It is associated with the depolymerization and rearrangement of cell components, including pectin, hemicellulose, cellulose, and other structural (glyco)proteins. Expansins are structural proteins to loosen plant cell wall polysaccharides in a pH-dependent manner and play pivotal roles in the process of fruit development, ripening, and softening. Rubus chingii Hu, a unique Chinese red raspberry, is a prestigious pharmaceutical and nutraceutical dual-function food with great economic value. Thirty-three RchEXPs were predicted by genome-wide identification in this study, containing twenty-seven α-expansins (EXPAs), three β-expansins (EXPBs), one expansin-like A (EXPLA), and two expansin-like B (EXPLBs). Subsequently, molecular characteristics, gene structure and motif compositions, phylogenetic relationships, chromosomal location, collinearity, and regulatory elements were further profiled. Furthermore, transcriptome sequencing (RNA-seq) and real-time quantitative PCR assays of fruits from different developmental stages and lineages showed that the group of RchEXPA5, RchEXPA7, and RchEXPA15 were synergistically involved in fruit expanding and ripening, while another group of RchEXPA6 and RchEXPA26 might be essential for fruit ripening and softening. They were regulated by both abscisic acid and ethylene and were collinear with phylogenetic relationships in the same group. Our new findings laid the molecular foundation for improving the fruit texture and shelf life of R. chingii medicinal and edible fruit.

Genome-wide identification and transcriptome profiling expression analysis of the U-box E3 ubiquitin ligase gene family related to abiotic stress in maize (Zea mays L.)

BackgroundThe U-box gene family encodes E3 ubiquitin ligases involved in plant hormone signaling pathways and abiotic stress responses. However, there has yet to be a comprehensive analysis of the U-box gene family in maize (Zea mays L.) and its responses to abiotic stress.ResultsIn this study, 85 U-box family proteins were identified in maize and were classified into four subfamilies based on phylogenetic analysis. In addition to the conserved U-box domain, we identified additional functional domains, including Pkinase, ARM, KAP and Tyr domains, by analyzing the conserved motifs and gene structures. Chromosomal localization and collinearity analysis revealed that gene duplications may have contributed to the expansion and evolution of the U-box gene family. GO annotation and KEGG pathway enrichment analysis identified a total of 105 GO terms and 21 KEGG pathways that were notably enriched, including ubiquitin-protein transferase activity, ubiquitin conjugating enzyme activity and ubiquitin-mediated proteolysis pathway. Tissue expression analysis showed that some ZmPUB genes were specifically expressed in certain tissues and that this could be due to their functions. In addition, RNA-seq data for maize seedlings under salt stress revealed 16 stress-inducible plant U-box genes, of which 10 genes were upregulated and 6 genes were downregulated. The qRT-PCR results for genes responding to abiotic stress were consistent with the transcriptome analysis. Among them, ZmPUB13, ZmPUB18, ZmPUB19 and ZmPUB68 were upregulated under all three abiotic stress conditions. Subcellular localization analysis showed that ZmPUB19 and ZmPUB59 were located in the nucleus.ConclusionsOverall, our study provides a comprehensive analysis of the U-box gene family in maize and its responses to abiotic stress, suggesting that U-box genes play an important role in the stress response and providing insights into the regulatory mechanisms underlying the response to abiotic stress in maize.

Genome-Wide Bioinformatics Analysis of SWEET Gene Family and Expression Verification of Candidate PaSWEET Genes in Potentilla anserina

Sugars act as the main energy sources in many fruit and vegetable crops. The biosynthesis and transportation of sugars are crucial and especially contribute to growth and development. SWEET is an important gene family that plays a vital role in plants’ growth, development, and adaptation to various types of stresses (biotic and abiotic). Although SWEET genes have been identified in numerous plant species, there is no information on SWEETs in Potentilla anserina. In the present study, we performed a comprehensive genome-wide bioinformatics analysis and identified a total of 23 candidate PaSWEETs genes in the Potentilla anserina genome, which were randomly distributed on ten different chromosomes. The phylogenetic analysis, chromosomal location, gene structure, specific cis-elements, protein interaction network, and physiological characteristics of these genes were systematically examined. The identified results of the phylogenetic relationship with Arabidopsis thaliana revealed that these PaSWEET genes were divided into four clades (I, II, III, and IV). Moreover, tissue-specific gene expression through quantitative real-time polymerase chain reaction (qRT-PCR) validation exposed that the identified PaSWEETs were differentially expressed in various tissues (roots, stems, leaves, and flowers). Mainly, the relative fold gene expression in swollen and unswollen tubers effectively revealed that PaSWEETs (7, 9, and 12) were highly expressed (300-, 120-, and 100-fold) in swollen tubers. To further elucidate the function of PaSWEETs (7, 9, and 12), their subcellular location was confirmed by inserting them into tobacco leaves, and it was noted that these genes were present on the cell membrane. On the basis of the overall results, it is suggested that PaSWEETs (7, 9, and 12) are the candidate genes involved in swollen tuber formation in P. anserina. In crux, we speculated that our study provides a valuable theoretical base for further in-depth function analysis of the PaSWEET gene family and their role in tuber development and further enhancing the molecular breeding of Potentilla anserina.

Comprehensive analysis of annexin gene family and its expression in response to branching architecture and salt stress in crape myrtle

BackgroundAnnexin (ANN) is calcium (Ca2+)-dependent and phospholipid binding protein family, which is involved in plant growth and development and response to various stresses. However, little known about ANN genes were identified from crape myrtle, an ornamental horticultural plant widely cultivated in the world.ResultsHere, 9 LiANN genes were identified from Lagerstroemia indica, and their characterizations and functions were investigated in L. indica for the first time. The LiANN genes were divided into 2 subfamilies. The gene structure, chromosomal location, and collinearity relationship were also explored. In addition, the GO annotation analysis of these LiANNs indicated that they are enriched in molecular functions, cellular components, and biological processes. Moreover, transcription factors (TFs) prediction analysis revealed that bHLH, MYB, NAC, and other TFs can interact with the LiANN promoters. Interestingly, the LiANN2/4/6–9 were demonstrated to play critical roles in the branching architecture of crape myrtle. Furthermore, the LiANN2/6/8/9 were differentially expressed under salt treatment, and a series of TFs regulating LiANN2/6/8/9 expression were predicted to play essential roles in salt resistance.ConclusionsThese results shed light on profile and function of the LiANN gene family, and lay a foundation for further studies of the LiANN genes.

The Silencing of GhPIP5K2 and GhPIP5K22 Weakens Abiotic Stress Tolerance in Upland Cotton (Gossypium hirsutum).

Phosphatidylinositol 4-phosphate 5-kinases (PIP5Ks), essential enzymes in the phosphatidylinositol signaling pathway, are crucial for the abiotic stress responses and the overall growth and development of plants. However, the GhPIP5Ks had not been systematically studied, and their function in upland cotton was unknown. This study identified a total of 28 GhPIP5Ks, and determined their chromosomal locations, gene structures, protein motifs and cis-acting elements via bioinformatics analysis. A quantitative real-time PCR (qRT‒PCR) analysis showed that most GhPIP5Ks were upregulated under different stresses. A virus-induced gene silencing (VIGS) assay indicated that the superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) activities were significantly decreased, while malondialdehyde (MDA) content were significantly increased in GhPIP5K2- and GhPIP5K22-silenced upland cotton plants under abiotic stress. Furthermore, the expression of the stress marker genes GhHSFB2A, GhHSFB2B, GhDREB2A, GhDREB2C, GhRD20-1, GhRD29A, GhBIN2, GhCBL3, GhNHX1, GhPP2C, GhCBF1, GhSnRK2.6 and GhCIPK6 was significantly decreased in the silenced plants after exposure to stress. These results revealed that the silencing of GhPIP5K2 and GhPIP5K22 weakened the tolerance to abiotic stresses. These discoveries provide a foundation for further inquiry into the actions of the GhPIP5K gene family in regulating the response and resistance mechanisms of cotton to abiotic stresses.

Phylogenomic curation of Ovate Family Proteins (OFPs) in the U's Triangle of Brassica L. indicates stress-induced growth modulation.

The Ovate Family Proteins (OFPs) gene family houses a class of proteins that are involved in regulating plant growth and development. To date, there is no report of the simultaneous functional characterization of this gene family in all members of U's Triangle of Brassica. Here, we retrieved a combined total of 256 OFP protein sequences and analyzed their chromosomal localization, gene structure, conserved protein motif domains, and the pattern of cis-acting regulatory elements. The abundance of light-responsive elements like G-box, MRE, and GT1 motif suggests that OFPs are sensitive to the stimuli of light. The protein-protein interaction network analysis revealed that OFP05 and its orthologous genes were involved in regulating the process of transcriptional repression through their interaction with homeodomain transcription factors like KNAT and BLH. The presence of domains like DNA binding 2 and its superfamily speculated the involvement of OFPs in regulating gene expression. The biotic and abiotic stress, and the tissue-specific expression analysis of the RNA-seq datasets revealed that some of the genes such as BjuOFP30, and BnaOFP27, BolOFP11, and BolOFP10 were highly upregulated in seed coat at the mature stage and roots under various chemical stress conditions respectively which suggests their crucial role in plant growth and development processes. Experimental validation of prominent BnaOFPs such as BnaOFP27 confirmed their involvement in regulating gene expression under salinity, heavy metal, drought, heat, and cold stress. The GO and KEGG pathway enrichment analysis also sheds light on the involvement of OFPs in regulating plant growth and development. These findings have the potential to serve as a forerunner for future studies in terms of functionally diverse analysis of the OFP gene family in Brassica and other plant species.

Identification and expression analysis of citrate synthase 3 gene family members in apple

As one of the key enzymes in cell metabolism, the activity of citrate synthase 3 (CS3) regulates the substance and energy metabolism of organisms. The protein members of CS3 family were identified from the whole genome of apple, and bioinformatics analysis was performed and expression patterns were analyzed to provide a theoretical basis for studying the potential function of CS3 gene in apple. BLASTp was used to identify members of the apple CS3 family based on the GDR database, and the basic information of CS3 protein sequence, subcellular localization, domain composition, phylogenetic relationship and chromosome localization were analyzed by Pfam, SMART, MEGA5.0, clustalx.exe, ExPASy Proteomics Server, MEGAX, SOPMA, MEME, WoLF PSORT and other software. The tissue expression and inducible expression characteristics of 6 CS3 genes in apple were determined by acid content and real-time fluorescence quantitative polymerase chain reaction (qRT-PCR). Apple CS3 gene family contains 6 members, and these CS3 proteins contain 473-608 amino acid residues, with isoelectric point distribution between 7.21 and 8.82. Subcellular localization results showed that CS3 protein was located in mitochondria and chloroplasts, respectively. Phylogenetic analysis divided them into 3 categories, and the number of genes in each subfamily was 2. Chromosome localization analysis showed that CS3 gene was distributed on different chromosomes of apple. The secondary structure of protein is mainly α-helix, followed by random curling, and the proportion of β-angle is the smallest. The 6 members were all expressed in different apple tissues. The overall expression trend from high to low was the highest relative expression content of MdCS3.4, followed by MdCS3.6, and the relative expression level of other members was in the order of MdCS3.3 > MdCS3.2 > MdCS3.1 > MdCS3.5. qRT-PCR results showed that MdCS3.1 and MdCS3.3 genes had the highest relative expression in the pulp of 'Chengji No. 1' with low acid content, and MdCS3.2 and MdCS3.3 genes in the pulp of 'Asda' with higher acid content had the highest relative expression. Therefore, in this study, the relative expression of CS3 gene in apple cultivars with different acid content in different apple varieties was detected, and its role in apple fruit acid synthesis was analyzed. The experimental results showed that the relative expression of CS3 gene in different apple varieties was different, which provided a reference for the subsequent study of the quality formation mechanism of apple.

X Chromosome-Specific Repeats in Non-Domestic Bovidae.

Repetitive sequences form a substantial and still enigmatic part of the mammalian genome. We isolated repetitive DNA blocks of the X chromosomes of three species of the family Bovidae: Kobus defassa (KDEXr sequence), Bos taurus (BTAXr sequence) and Antilope cervicapra (ACEXr sequence). The copy numbers of the isolated sequences were assessed using qPCR, and their chromosomal localisations were analysed using FISH in ten bovid tribes and in outgroup species. Besides their localisation on the X chromosome, their presence was also revealed on the Y chromosome and autosomes in several species. The KDEXr sequence abundant in most Bovidae species also occurs in distant taxa (Perissodactyla and Carnivora) and seems to be evolutionarily older than BTAXr and ACEXr. The ACEXr sequence, visible only in several Antilopini species using FISH, is probably the youngest, and arised in an ancestor common to Bovidae and Cervidae. All three repetitive sequences analysed in this study are interspersed among gene-rich regions on the X chromosomes, apparently preventing the crossing-over in their close vicinity. This study demonstrates that repetitive sequences on the X chromosomes have undergone a fast evolution, and their variation among related species can be beneficial for evolutionary studies.

Genome-wide identification of MAPK family in papaya (Carica papaya) and their involvement in fruit postharvest ripening

BackgroundPapaya (Carica papaya) is an economically important fruit cultivated in the tropical and subtropical regions of China. However, the rapid softening rate after postharvest leads to a short shelf-life and considerable economic losses. Accordingly, understanding the mechanisms underlying fruit postharvest softening will be a reasonable way to maintain fruit quality and extend its shelf-life.ResultsMitogen-activated protein kinases (MAPKs) are conserved and play essential roles in response to biotic and abiotic stresses. However, the MAPK family remain poorly studied in papaya. Here, a total of nine putative CpMAPK members were identified within papaya genome, and a comprehensive genome-wide characterization of the CpMAPKs was performed, including evolutionary relationships, conserved domains, gene structures, chromosomal locations, cis-regulatory elements and expression profiles in response to phytohormone and antioxidant organic compound treatments during fruit postharvest ripening. Our findings showed that nearly all CpMAPKs harbored the conserved P-loop, C-loop and activation loop domains. Phylogenetic analysis showed that CpMAPK members could be categorized into four groups (A-D), with the members within the same groups displaying high similarity in protein domains and intron–exon organizations. Moreover, a number of cis-acting elements related to hormone signaling, circadian rhythm, or low-temperature stresses were identified in the promoters of CpMAPKs. Notably, gene expression profiles demonstrated that CpMAPKs exhibited various responses to 2-chloroethylphosphonic acid (ethephon), 1-methylcyclopropene (1-MCP) and the combined ascorbic acid (AsA) and chitosan (CTS) treatments during papaya postharvest ripening. Among them, both CpMAPK9 and CpMAPK20 displayed significant induction in papaya flesh by ethephon treatment, and were pronounced inhibition after AsA and CTS treatments at 16 d compared to those of natural ripening control, suggesting that they potentially involve in fruit postharvest ripening through ethylene signaling pathway or modulating cell wall metabolism.ConclusionThis study will provide some valuable insights into future functional characterization of CpMAPKs, and hold great potential for further understanding the molecular mechanisms underlying papaya fruit postharvest ripening.

Genome-wide identification of the expansin gene family in netted melon and their transcriptional responses to fruit peel cracking.

Fruit cracking not only affects the appearance of netted melons (Cucumis melo L. var. reticulatus Naud.) but also decreases their marketability. Herein, to comprehensively understand the role of expansin (EXP) proteins in netted melon, bioinformatics methods were employed to discover the EXP gene family in the melon genome and analyze its characteristic features. Furthermore, transcriptomics analysis was performed to determine the expression patterns of melon EXP (CmEXP) genes in crack-tolerant and crack-susceptible netted melon varieties. Thirty-three CmEXP genes were identified. Chromosomal location analysis revealed that CmEXP gene distribution was uneven on 12 chromosomes. In addition, phylogenetic tree analysis revealed that CmEXP genes could be categorized into four subgroups, among which the EXPA subgroup had the most members. The same subgroup members shared similar protein motifs and gene structures. Thirteen duplicate events were identified in the 33 CmEXP genes. Collinearity analysis revealed that the CmEXP genes had 50, 50, and 44 orthologous genes with EXP genes in cucumber, watermelon, and Arabidopsis, respectively. However, only nine orthologous EXP genes were observed in rice. Promoter cis-acting element analysis demonstrated that numerous cis-acting elements in the upstream promoter region of CmEXP genes participate in plant growth, development, and environmental stress responses. Transcriptomics analysis revealed 14 differentially expressed genes (DEGs) in the non-cracked fruit peels between the crack-tolerant variety 'Xizhoumi 17' (N17) and the crack-susceptible variety 'Xizhoumi 25' (N25). Among the 14 genes, 11 were upregulated, whereas the remaining three were downregulated in N17. In the non-cracked (N25) and cracked (C25) fruit peels of 'Xizhoumi 25', 24 DEGs were identified, and 4 of them were upregulated, whereas the remaining 20 were downregulated in N25. In the two datasets, only CmEXPB1 exhibited consistently upregulated expression, indicating its importance in the fruit peel crack resistance of netted melon. Transcription factor prediction revealed 56 potential transcription factors that regulate CmEXPB1 expression. Our study findings enrich the understanding of the CmEXP gene family and present candidate genes for the molecular breeding of fruit peel crack resistance of netted melon.