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Abstract The color of red-skinned pear (Pyrus spp.) is primarily attributed to accumulation of anthocyanins, which provide nutritional benefits for human health and are closely associated with the commercial value of fruits. Here, we reported the functional characterization of a R2R3-MYB repressor PyMYB107, which forms an ‘activator-repressor’ loop to control anthocyanin accumulation in the red-skinned pear. PyMYB107 overexpression inhibited anthocyanin biosynthesis in both pear calli and fruits, while virus-induced gene silencing of PyMYB107 increased anthocyanin accumulation in pear fruits. Furthermore, ectopic expression of PyMYB107 decreased anthocyanin accumulation in tomato, strawberry and tobacco. PyMYB107 can competitively bind to PybHLH3 with PyMYB10/MYB114, thereby suppressing the transcriptional activation of key anthocyanin biosynthesis genes, PyANS and PyUFGT. Site-directed mutagenesis showed that mutations within the R3 domain and EAR motif of PyMYB107 eliminated its repressive activity. Additionally, PyMYB107 exhibited a comparable expression pattern to PyMYB10/MYB114 and was transcriptionally activated by them. Our finding advanced comprehension of the repression mechanism underlying anthocyanin accumulation, providing valuable molecular insights into improving quality of pear fruits.

Abstract Background Minimally invasive plate osteosynthesis (MIPO) via percutaneous plate placement on the distal medial tibia can be performed with minimizes soft tissue injury and produces good clinical results. However, the difficulty with MIPO lies in how to achieve satisfactory fracture reduction and maintain that reduction via indirect reduction techniques to facilitate internal fixation. The purpose of this study was to compare the effects of AO distractor and manual traction reduction techniques combined with MIPO in the treatment of distal tibia fractures. Methods Between January 2013 and December 2019, 58 patients with a distal tibia fracture were treated using MIPO. Patients were divided into two groups according to the indirect reduction method that was used: 26 patients were reduced with manual traction(group M), and 32 were reduced with an AO distractor (group A).Time until union and clinical outcomes including AOFAS ankle-rating score and ankle range of ankle motion at final follow-up were compared. Mean operative time, incision length, blood loss and postoperative complications were recorded via chart review. Radiographic results at final follow-up were assessed for tibial angulation and shortening by a blinded reader. Results Mean operative time, incision length, and blood loss in group A were significantly lower than in group M(p = 0.019, 0.018 and 0.016, respectively).Radiographic evidence of bony union was seen in all cases, and mean time until union was equivalent between the two groups (p = 0.384).Skin irritation was noted in one case(3.1%) in group A and three cases(11.5%)in group M, but the symptoms were not severe and the plate was removed after bony union. There was no statistically significant difference in postoperative complications between the two groups(p = 0.461). Mean AOFAS score and range of ankle motion were equivalent between the two groups, as were varus deformity, valgus deformity, anterior angulation and posterior angulation. No patients had gross angular deformity. Mean tibial shortening was not significantly different between the two groups, and no patients had tibial shortening > 10 mm. Conclusion Both an AO distractor and manual traction reduction techniques prior to MIPO in the treatment of distal tibial fractures permit a high fracture healing rate and satisfying functional outcomes with few wound healing complications. An AO distractor is an excellent indirect reduction method that may improve operative efficiency and reduce the risk of soft tissue injury.

Abstract Background Structural variants (SVs) play important roles in adaptation evolution and species diversification. Especially, in plants, many phenotypes of response to the environment were found to be associated with SVs. Despite the prevalence and significance of SVs, long insertions remain poorly detected and studied in all but model species. Results We used whole-genome resequencing of paired reads from 80 Asian butternuts to detect long insertions and further analyse their characteristics and potential functional effects. By combining of mapping-based and de novo assembly-based methods, we obtained a multiple related species pangenome representing higher taxonomic groups. We obtained 89,312 distinct contigs totaling 147,773,999 base pair (bp) of new sequences, of which 347 were putative long insertions placed in the reference genome. Most of the putative long insertions appeared in multiple species; in contrast, only 62 putative long insertions appeared in one species, which may be involved in the response to the environment. 65 putative long insertions fell into 61 distinct protein-coding genes involved in plant development, and 105 putative long insertions fell into upstream of 106 distinct protein-coding genes involved in cellular respiration. 3,367 genes were annotated in 2,606 contigs. We propose PLAINS ( https://github.com/CMB-BNU/PLAINS.git ), a streamlined, comprehensive pipeline for the prediction and analysis of long insertions using whole-genome resequencing. Conclusions Our study lays down an important foundation for further whole-genome long insertion studies, allowing the investigation of their effects by experiments.

Abstract Background Alternative splicing (AS) is an important mechanism of posttranscriptional modification and dynamically regulates multiple physiological processes in plants, including fruit ripening. However, little is known about alternative splicing during fruit development in fleshy fruits. Results We studied the alternative splicing at the immature and ripe stages during fruit development in cucumber, melon, papaya and peach. We found that 14.96–17.48% of multiexon genes exhibited alternative splicing. Intron retention was not always the most frequent event, indicating that the alternative splicing pattern during different developmental process differs. Alternative splicing was significantly more prevalent at the ripe stage than at the immature stage in cucumber and melon, while the opposite trend was shown in papaya and peach, implying that developmental stages adopt different alternative splicing strategies for their specific functions. Some genes involved in fruit ripening underwent stage-specific alternative splicing, indicating that alternative splicing regulates fruits ripening. Conserved alternative splicing events did not appear to be stage-specific. Clustering fruit developmental stages across the four species based on alternative splicing profiles resulted in species-specific clustering, suggesting that diversification of alternative splicing contributes to lineage-specific evolution in fleshy fruits. Conclusions We obtained high quality transcriptomes and alternative splicing events during fruit development across the four species. Dynamics and nonconserved alternative splicing were discovered. The candidate stage-specific AS genes involved in fruit ripening will provide valuable insight into the roles of alternative splicing during the developmental processes of fleshy fruits.

Abstract Background With the rapid development of accurate sequencing and assembly technologies, an increasing number of high-quality chromosome-level and haplotype-resolved assemblies of genomic sequences have been derived, from which there will be great opportunities for computational pangenomics. Although genome graphs are among the most useful models for pangenome representation, their structural complexity makes it difficult to present genome information intuitively, such as the linear reference genome. Thus, efficiently and accurately analyzing the genome graph spatial structure and coordinating the information remains a substantial challenge. Results We developed a new method, a colored superbubble (cSupB), that can overcome the complexity of graphs and organize a set of species- or population-specific haplotype sequences of interest. Based on this model, we propose a tri-tuple coordinate system that combines an offset value, topological structure and sample information. Additionally, cSupB provides a novel method that utilizes complete topological information and efficiently detects small indels (

Abstract Background Alternative splicing (AS) plays a critical regulatory role in modulating transcriptome and proteome diversity. In particular, it increases the functional diversity of proteins. Recent genome-wide analysis of AS using RNA-Seq has revealed that AS is highly pervasive in plants. Furthermore, it has been suggested that most AS events are subject to tissue-specific regulation. Description To reveal the functional characteristics induced by AS and tissue-specific splicing events, a database for exploring these characteristics is needed, especially in plants. To address these goals, we constructed a database of annotated transcripts generated by alternative splicing in cucumbers (CuAS: http://cmb.bnu.edu.cn/alt_iso/index.php ) that integrates genomic annotations, isoform-level functions, isoform-level features, and tissue-specific AS events among multiple tissues. CuAS supports a retrieval system that identifies unique IDs (gene ID, isoform ID, UniProt ID, and gene name), chromosomal positions, and gene families, and a browser for visualization of each gene. Conclusion We believe that CuAS could be helpful for revealing the novel functional characteristics induced by AS and tissue-specific AS events in cucumbers. CuAS is freely available at http://cmb.bnu.edu.cn/alt_iso/index.php .

Abstract Background Temperature is a major determinant of spontaneous mutation, but the precise mode, and the underlying mechanisms, of the temperature influences remain less clear. Here we used a mutation accumulation approach combined with whole-genome sequencing to investigate the temperature dependence of spontaneous mutation in an Escherichia coli strain. Experiments were performed under aerobic conditions at 25, 28 and 37 °C, three temperatures that were non-stressful for the bacterium but caused significantly different bacterial growth rates. Results Mutation rate did not differ between 25 and 28 °C, but was higher at 37 °C. Detailed analyses of the molecular spectrum of mutations were performed; and a particularly interesting finding is that higher temperature led to a bias of mutation to coding, relative to noncoding, DNA. Furthermore, the temperature response of mutation rate was extremely similar to that of metabolic rate, consistent with an idea that metabolic rate predicts mutation rate. Conclusions Temperature affects mutation rate and the types of mutation supply, both being crucial for the opportunity of natural selection. Our results help understand how temperature drives evolutionary speed of organisms and thus the global patterns of biodiversity. This study also lend support to the metabolic theory of ecology for linking metabolic rate and molecular evolution rate.

Abstract Background Most published genome sequences are drafts, and most are dominated by computational gene prediction. Draft genomes typically incorporate considerable sequence data that are not assigned to chromosomes, and predicted genes without quality confidence measures. The current Actinidia chinensis (kiwifruit) ‘Hongyang’ draft genome has 164 Mb of sequences unassigned to pseudo-chromosomes, and omissions have been identified in the gene models. Results A second genome of an A. chinensis (genotype Red5) was fully sequenced. This new sequence resulted in a 554.0 Mb assembly with all but 6 Mb assigned to pseudo-chromosomes. Pseudo-chromosomal comparisons showed a considerable number of translocation events have occurred following a whole genome duplication (WGD) event some consistent with centromeric Robertsonian-like translocations. RNA sequencing data from 12 tissues and ab initio analysis informed a genome-wide manual annotation, using the WebApollo tool. In total, 33,044 gene loci represented by 33,123 isoforms were identified, named and tagged for quality of evidential support. Of these 3114 (9.4%) were identical to a protein within ‘Hongyang’ The Kiwifruit Information Resource (KIR v2). Some proportion of the differences will be varietal polymorphisms. However, as most computationally predicted Red5 models required manual re-annotation this proportion is expected to be small. The quality of the new gene models was tested by fully sequencing 550 cloned ‘Hort16A’ cDNAs and comparing with the predicted protein models for Red5 and both the original ‘Hongyang’ assembly and the revised annotation from KIR v2. Only 48.9% and 63.5% of the cDNAs had a match with 90% identity or better to the original and revised ‘Hongyang’ annotation, respectively, compared with 90.9% to the Red5 models. Conclusions Our study highlights the need to take a cautious approach to draft genomes and computationally predicted genes. Our use of the manual annotation tool WebApollo facilitated manual checking and correction of gene models enabling improvement of computational prediction. This utility was especially relevant for certain types of gene families such as the EXPANSIN like genes. Finally, this high quality gene set will supply the kiwifruit and general plant community with a new tool for genomics and other comparative analysis.

Abstract Background Alternative splicing (AS) is an important post-transcriptional process. It has been suggested that most AS events are subject to tissue-specific regulation. However, the global dynamics of AS in different tissues are poorly explored. Results To analyse global changes in AS in multiple tissues, we identified the AS events and constructed a comprehensive catalogue of AS events within each tissue based on the genome-wide RNA-seq reads from ten tissues in cucumber. First, we found that 58% of the multi-exon genes underwent AS. We further obtained 565 genes with significantly more AS events compared with random genes. These genes were found significant enrichment in biological processes related to the regulation of actin filament length. Second, significantly different AS event profiles among ten tissues were found. The tissues with the same origin of development are more likely to have a relatively similar AS profile. Moreover, 7370 genes showed tissue-specific AS events and were highly enriched in biological processes related to the positive regulation of cellular component organization. Root-specificity AS genes were related to the cellular response to DNA damage stimulus. Third, the genes with different intron retention (IR) patterns among the ten tissues showed significant difference in GC percentages of the retained intron, and the number of exons and FPKM of the major transcripts. Conclusions Our study provided a comprehensive view of AS in multiple tissues. We revealed novel insights into the patterns of AS in multiple tissues and the tissue-specific AS in cucumber.

Abstract Background Because phylogenetic inference is an important basis for answering many evolutionary problems, a large number of algorithms have been developed. Some of these algorithms have been improved by integrating gene evolution models with the expectation of accommodating the hierarchy of evolutionary processes. To the best of our knowledge, however, there still is no single unifying model or algorithm that can take all evolutionary processes into account through a stepwise or simultaneous method. Results On the basis of three existing phylogenetic inference algorithms, we built an integrated pipeline for inferring the evolutionary history of a given gene family; this pipeline can model gene sequence evolution, gene duplication-loss, gene transfer and multispecies coalescent processes. As a case study, we applied this pipeline to the STIMATE (TMEM110) gene family, which has recently been reported to play an important role in store-operated Ca2+ entry (SOCE) mediated by ORAI and STIM proteins. We inferred their phylogenetic trees in 69 sequenced chordate genomes. Conclusions By integrating three tree reconstruction algorithms with diverse evolutionary models, a pipeline for inferring the evolutionary history of a gene family was developed, and its application was demonstrated.

Abstract Background Chinese primary care settings have a heavy patient load, shortage of physicians, limited medical resources and low medical literacy, making it difficult to screen for developmental disorders in infants. The Infant Neurological International Battery (INFANIB) for the assessment of neuromotor developmental disorders in infants aged 0 ~ 18 months is widely applied in community health service centers because of its simplicity, time-saving advantages and short learning curve. We aimed to develop and assess a Chinese version of the INFANIB. Methods A Chinese version of the INFANIB was developed. Fifty-five preterm and 49 full-term infants with high risk of neurodevelopmental delays were assessed using the Chinese version of the INFANIB at 3, 7 and 10 months after birth. The Peabody Developmental Motor Scale (PDMS) was simultaneously used to assess the children with abnormalities and diagnose cerebral palsy. The sensitivity, specificity, positive predictive value and negative predictive value of the scale were calculated. Results At birth, a higher proportion of full-term infants had asphyxia (p p = 0.003) and hyperbilirubinemia ( p = 0.022). The interclass correlation coefficient and intraclass correlation coefficient values for the INFANIB at 3, 7 and 10 months were >0.8, indicating excellent reliability with regard to inter- and intraobserver differences. The specificity, sensitivity, positive predictive value and negative predictive value were high for both high-risk premature infants and full-term infants at the age of 10 months. For premature infants at the age of 7 months or below, INFANIB had low validity for detecting abnormalities. Conclusions The Chinese version of the INFANIB can be useful for screening infants with high-risk for neuromotor abnormality in Chinese primary care settings.