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Insertions and deletions constitute the second most important source of natural genomic variation. Insertions and deletions make up to 25% of genomic variants in humans and are involved in complex evolutionary processes including genomic rearrangements, adaptation, and speciation. Recent advances in long-read sequencing technologies allow detailed inference of insertions and deletion variation in species and populations. Yet, despite their importance, evolutionary studies have traditionally ignored or mishandled insertions and deletions due to a lack of comprehensive methodologies and statistical models of insertions and deletion dynamics. Here, we discuss methods for describing insertions and deletion variation and modeling insertions and deletions over evolutionary time. We provide practical advice for tackling insertions and deletions in genomic sequences and illustrate our discussion with examples of insertions and deletion-induced effects in human and other natural populations and their contribution to evolutionary processes. We outline promising directions for future developments in statistical methodologies that would allow researchers to analyze insertions and deletion variation and their effects in large genomic data sets and to incorporate insertions and deletions in evolutionary inference. [ABSTRACT FROM AUTHOR]

Background: Variant interpretation is essential for identifying patients' disease-causing genetic variants amongst the millions detected in their genomes. Hundreds of Variant Impact Predictors (VIPs), also known as Variant Effect Predictors (VEPs), have been developed for this purpose, with a variety of methodologies and goals. To facilitate the exploration of available VIP options, we have created the Variant Impact Predictor database (VIPdb). Results: The Variant Impact Predictor database (VIPdb) version 2 presents a collection of VIPs developed over the past three decades, summarizing their characteristics, ClinGen calibrated scores, CAGI assessment results, publication details, access information, and citation patterns. We previously summarized 217 VIPs and their features in VIPdb in 2019. Building upon this foundation, we identified and categorized an additional 190 VIPs, resulting in a total of 407 VIPs in VIPdb version 2. The majority of the VIPs have the capacity to predict the impacts of single nucleotide variants and nonsynonymous variants. More VIPs tailored to predict the impacts of insertions and deletions have been developed since the 2010s. In contrast, relatively few VIPs are dedicated to the prediction of splicing, structural, synonymous, and regulatory variants. The increasing rate of citations to VIPs reflects the ongoing growth in their use, and the evolving trends in citations reveal development in the field and individual methods. Conclusions: VIPdb version 2 summarizes 407 VIPs and their features, potentially facilitating VIP exploration for various variant interpretation applications. VIPdb is available at https://genomeinterpretation.org/vipdb [ABSTRACT FROM AUTHOR]

Background: White lupin (Lupinus albus L.) is a high-protein Old World grain legume with remarkable food and feed production interest. It is sown in autumn or early spring, depending on the local agroclimatic conditions. This study aimed to identify allelic variants associated with vernalization responsiveness, in order to improve our knowledge of legume flowering regulatory pathways and develop molecular selection tools for the desired phenology as required for current breeding and adaptation to the changing climate. Results: Some 120 white lupin accessions originating from a wide range of environments of Europe, Africa, and Asia were phenotyped under field conditions in three environments with different intensities of vernalization, namely, a Mediterranean and a subcontinental climate sites of Italy under autumn sowing, and a suboceanic climate site of France under spring sowing. Two hundred sixty-two individual genotypes extracted from them were phenotyped in a greenhouse under long-day photoperiod without vernalization. Phenology data, and marker data generated by Diversity Arrays Technology sequencing (DArT-seq) and by PCR-based screening targeting published quantitative trait loci (QTLs) from linkage map and newly identified insertion/deletion polymorphisms in the promoter region of the FLOWERING LOCUS T homolog, LalbFTc1 gene (Lalb_Chr14g0364281), were subjected to a genome-wide association study (GWAS). Population structure followed differences in phenology and isolation by distance pattern. The GWAS highlighted numerous loci significantly associated with flowering time, including four LalbFTc1 gene promoter deletions: 2388 bp and 2126 bp deletions at the 5' end, a 264 bp deletion in the middle and a 28 bp deletion at the 3' end of the promoter. Besides LalbFTc1 deletions, this set contained DArT-seq markers that matched previously published major QTLs in chromosomes Lalb_Chr02, Lalb_Chr13 and Lalb_Chr16, and newly discovered QTLs in other chromosomes. Conclusions: This study highlighted novel QTLs for flowering time and validated those already published, thereby providing novel evidence on the convergence of FTc1 gene functional evolution into the vernalization pathway in Old World lupin species. Moreover, this research provided the set of loci specific for extreme phenotypes (the earliest or the latest) awaiting further implementation in marker-assisted selection for spring- or winter sowing. [ABSTRACT FROM AUTHOR]

Deficiencies in DNA mismatch repair (MMRd) leave characteristic footprints of microsatellite instability (MSI) in cancer genomes. We used data from the Cancer Genome Atlas and International Cancer Genome Consortium to conduct a comprehensive analysis of MSI-associated cancers, focusing on indel mutational signatures. We classified MSI-high genomes into two subtypes based on their indel profiles: deletion-dominant (MMRd-del) and insertion-dominant (MMRd-ins). Compared with MMRd-del genomes, MMRd-ins genomes exhibit distinct mutational and transcriptomic features, including a higher prevalence of T>C substitutions and related mutation signatures. Short insertions and deletions in MMRd-ins and MMRd-del genomes target different sets of genes, resulting in distinct indel profiles between the two subtypes. In addition, indels in the MMRd-ins genomes are enriched with subclonal alterations that provide clues about a distinct evolutionary relationship between the MMRd-ins and MMRd-del genomes. Notably, the transcriptome analysis indicated that MMRd-ins cancers upregulate immune-related genes, show a high level of immune cell infiltration, and display an elevated neoantigen burden. The genomic and transcriptomic distinctions between the two types of MMRd genomes highlight the heterogeneity of genetic mechanisms and resulting genomic footprints and transcriptomic changes in cancers, which has potential clinical implications. [ABSTRACT FROM AUTHOR]

Stathmin 1 (stmn1) gene, which is involved in the control of cell cycle progression, cell proliferation, differentiation and motility, is a crucial relay in the signal transduction of multiple signaling pathways. Reports on the function of stmn1 in fish are scarce. In this study, the characterization and function of stmn1a gene were explored in an important food fish, namely bighead carp (Hypophthalmichthys nobilis). The bighead carp stmn1a (Hynstmn1a) genomic sequence, which has 2995 bp with an ORF of 447 bp, is evolutionarily conserved in teleost fish. The highest level of expression of Hynstmn1a is in the un-fertilized eggs and spleen of adult bighead carp. The overexpression of Hynstmn1a facilitates cell cycle progression and suppresses apoptosis in EPC cells. In an siRNA assay, disruption of stmn1a in EPC cells resulted in the inhibition of cell cycle progression and promotion of apoptosis. Analysis of promoter activity indicated that an upstream transcription factor (foxo4) regulates Hynstmn1a. Zebrafish mutants with stmn1a knockout by CRISPR/Cas9 exhibited a significant retardation of body length (p < 0.05). There was no significant difference in body weight between stmn1a-mutant zebrafish and wildtype (p = 0.053). A 22-bp Indel in the promoter region of Hynstmn1a with an observed heterozygosity (Ho) of 0.136 in a bighead carp population (n = 191). The polymorphism of this 22-bp Indel genotype is significantly associated with body length (BL), head length (HL) (p < 0.05), body weight (BW), head height (HH), and head width (HW) (p < 0.01), showing its genetic effects on growth promotion. This study sheds light on the function of stmn1a in fish and demonstrates the potential of the 22-bp Indel of Hynstmn1a as a gene marker when selecting breeding individuals for faster growth and a bigger head of bighead carp in aquaculture. [ABSTRACT FROM AUTHOR]

Maize(Zea mays. L) is a globally important crop, and understanding its genetic diversity is crucial for plant breeding phylogenetic analyses and comparative genetics. While nuclear markers have been extensively used for mapping agriculturally important genes, they are limited in recognizing characteristics, such as cytoplasmic male sterility and reciprocal cross hybrids. In this study, we performed next-generation sequencing of 176samples, and the maize cultivars represented five distinct groups. A total of 89 single nucleotide polymorphisms (SNPs) and 11 insertion/deletion polymorphisms (InDels) were identified. To enable high-throughput detection, we successfully amplified and confirmed 49 SNP and InDel markers, which were defined as a Varietal Chloroplast Panel (VCP) using the Kompetitive Allele Specific PCR (KASP). The specific markers provided a valuable tool for identifying chloroplast groups. The verification experiment, focusing on the identification of reciprocal cross hybrids and cytoplasmic male sterility hybrids, demonstrated the significant advantages of VCP markers in maternal inheritance characterization. Furthermore, only a small subset of these markers is needed to provide useful information, showcasing the effectiveness of these markers in elucidating the artificial selection process of elite maize lines. [ABSTRACT FROM AUTHOR]

We previously found that OsCUL3c is involved in the salt stress response. However, there are no definitive reports on the diversity of OsCUL3c in local Thai rice. In this study, we showed that the CUL3 group was clearly separated from the other CUL groups; next, we focused on OsCUL3c, the third CUL3 of the CUL3 family in rice, which is absent in Arabidopsis. A total of 111 SNPs and 28 indels over the OsCUL3c region, representing 79 haplotypes (haps), were found. Haplotyping revealed that group I (hap A and hap C) and group II (hap B1 and hap D) were different mutated variants, which showed their association with phenotypes under salt stress. These results were supported by cis-regulatory elements (CREs) and transcription factor binding sites (TFBSs) analyses. We found that LTR, MYC, [AP2; ERF], and NF-YB, which are related to salt stress, drought stress, and the response to abscisic acid (ABA), have distinct positions and numbers in the haplotypes of group I and group II. An RNA Seq analysis of the two predominant haplotypes from each group showed that the OsCUL3c expression of the group I representative was upregulated and that of group II was downregulated, which was confirmed by RT-qPCR. Promoter changes might affect the transcriptional responses to salt stress, leading to different regulatory mechanisms for the expression of different haplotypes. We speculate that OsCUL3c influences the regulation of salt-related responses, and haplotype variations play a role in this regulation. [ABSTRACT FROM AUTHOR]

An insertion/deletion (InDel) polymorphism study of PR/SET domain family 6 (PRDM6), myostatin (MSTN) and insulin-like growth factor 2 mRNA binding protein 1 (IGF2BP1) genes was conducted in Malabari and Attappady black goats. An association study of identified InDels and body measurement traits was also performed. Body measurements included body length, chest diameter, chest depth, canon circumference, hip width, and hip height at the hip cross. The body trunk index, the body length index, the canon circumference index, and the chest width index were calculated. The Hardy-Weinberg equilibrium (HWE) was tested using a Chi-square test. The observed heterozygosity (Ho), expected heterozygosity (He), and polymorphism information content (PIC) were calculated. A significant difference in body measurements was found across breeds, ages, and breed x age interactions. The PRDM6 InDel was also associated with body measurement traits, such as body height, canon circumference and canon circumference index. In both Malabari and Attappadi black MSTN and PRDM6 InDels were in a state of HWE, while IGF2BP1 InDels were not. Indel markers found in the present study may be used for marker-assisted selection of growth traits among goats. [ABSTRACT FROM AUTHOR]

Grifola frondosa is a valuable edible fungus with high nutritional and medicinal values. The mating systems of fungi not only offer practical strategies for breeding, but also have far-reaching effects on genetic variability. Grifola frondosa has been considered as a sexual species with a tetrapolar mating system based on little experimental data. In the present study, one group of test crosses and six groups of three-round mating experiments from two parental strains were conducted to determine the mating system in G. frondosa. A chi-squared test of the results of the test-cross mating experiments indicated that they satisfied Mendelian segregation, while a series of three-round mating experiments showed that Mendelian segregation was not satisfied, implying a segregation distortion phenomenon in G. frondosa. A genomic map of the G. frondosa strain, y59, grown from an LMCZ basidiospore, with 40.54 Mb and 12 chromosomes, was generated using genome, transcriptome and Hi-C sequencing technology. Based on the genomic annotation of G. frondosa, the mating-type loci A and B were located on chromosomes 1 and 11, respectively. The mating-type locus A coded for the β-fg protein, HD1, HD2 and MIP, in that order. The mating-type locus B consisted of six pheromone receptors (PRs) and five pheromone precursors (PPs) in a crossed order. Moreover, both HD and PR loci may have only one sublocus that determines the mating type in G. frondosa. The nonsynonymous SNP and indel mutations between the A1B1 and A2B2 mating-type strains and the reference genome of y59 only occurred on genes HD2 and PR1/2, preliminarily confirming that the mating type of the y59 strain was A1B2 and not A1B1. Based on the genetic evidence and the more reliable molecular evidence, the results reveal that the mating system of G. frondosa is tetrapolar. This study has important implications for the genetics and hybrid breeding of G. frondosa. [ABSTRACT FROM AUTHOR]

Sorghum peduncle length is one of the key agronomic traits in sorghum production and has a critical impact on sorghum mechanization for harvesting. In this study, an F2 segregating population was constructed with significantly different long-peduncle KY133B and short-peduncle KY123B as parents, and the sorghum peduncle length gene was preliminarily located using BSA-seq technology. The association analysis was performed by ED algorithm and SNP-index (or InDel-index) algorithm, and finally the sorghum peduncle length association region was targeted on chromosome 7 and 10. In this study, the BSA-seq technique was used to rapidly and efficiently locate key genetic segments of sorghum peduncle length, which laid the foundation for subsequent functional validation and molecular studies of peduncle length genes. [ABSTRACT FROM AUTHOR]

Background : Single nucleotide polymorphisms (SNPs) and insertions/deletions (InDels) are the most abundant genetic variations and widely distribute across the genomes in plant. Development of SNP and InDel markers is a valuable tool for genetics and genomic research in radish (Raphanus sativus L.). Results : In this study, a total of 366,679 single nucleotide polymorphisms (SNPs) and 97,973 insertion-deletion (InDel) markers were identified based on genome resequencing between 'YZH' and 'XHT'. In all, 53,343 SNPs and 4,257 InDels were detected in two cultivars by transcriptome sequencing. Among the InDel variations, 85 genomic and 15 transcriptomic InDels were newly developed and validated PCR. The 100 polymorphic InDels markers generated 207 alleles among 200 Chinese radish germplasm, with an average 2.07 of the number of alleles (Na) and with an average 0.33 of the polymorphism information content (PIC). Population structure and phylogenetic relationship revealed that the radish cultivars from northern China were clustered together and the southwest China cultivars were clustered together. RNA-Seq analysis revealed that 11,003 differentially expressed genes (DEGs) were identified between the two cultivars, of which 5,020 were upregulated and 5,983 were downregulated. In total, 145 flowering time-related DGEs were detected, most of which were involved in flowering time integrator, circadian clock/photoperiod autonomous, and vernalization pathways. In flowering time-related DGEs region, 150 transcriptomic SNPs and 9 InDels were obtained. Conclusions: The large amount of SNPs and InDels identified in this study will provide a valuable marker resource for radish genetic and genomic studies. The SNPs and InDels within flowering time-related DGEs provide fundamental insight into for dissecting molecular mechanism of bolting and flowering in radish. [ABSTRACT FROM AUTHOR]

The domestic canary (Serinus canaria) is one of the most common pet birds and has been extensively selected and bred over the last few centuries to constitute many different varieties. Plumage pigmentation is one of the main phenotypic traits that distinguish canary breeds and lines. Feather colours in these birds, similarly to other avian species, are mainly depended on the presence of two major types of pigments: carotenoids and melanins. In this study, we exploited whole genome sequencing (WGS) datasets produced from five canary lines or populations (Black Frosted Yellow, Opal, Onyx, Opal × Onyx and Mogno, some of which carrying different putative dilute alleles), complemented with other WGS datasets retrieved from previous studies, to identify candidate genes that might explain pigmentation variability across canary breeds and varieties. Sequencing data were obtained using a DNA pool‐seq approach and genomic data were compared using window‐based FST analyses. We identified signatures of selection in genomic regions harbouring genes involved in carotenoid‐derived pigmentation variants (CYP2J19, EDC, BCO2 and SCARB1), confirming the results reported by previous works, and identified several other signatures of selection in the correspondence of melanogenesis‐related genes (AGRP, ASIP, DCT, EDNRB, KITLG, MITF, MLPH, SLC45A2, TYRP1 and ZEB2). Two putative causative mutations were identified in the MLPH gene that may explain the Opal and Onyx dilute mutant alleles. Other signatures of selection were also identified that might explain additional phenotypic differences between the investigated canary populations. [ABSTRACT FROM AUTHOR]

Cytoplasmic male sterility (CMS) is predominantly used for F1 hybrid breeding and seed production in Sorghum. DNA markers to distinguish between normal fertile (CMS-N) and sterile (CMS-S) male cytoplasm can facilitate F1 hybrid cultivar development in Sorghum breeding programs. In this study, the complete chloroplast (cp) genome sequences of CMS-S and Korean Sorghum cultivars were obtained using next-generation sequencing. The de novo assembled genome size of ATx623, the CMS-S line of the chloroplast, was 140,644bp. When compared to the CMS-S and CMS-N cp genomes, 19 single nucleotide polymorphisms (SNPs) and 142 insertions and deletions (InDels) were identified, which can be used for marker development for breeding, population genetics, and evolution studies. Two InDel markers with sizes greater than 20 bp were developed to distinguish cytotypes based on the copy number variation of lengths as 28 and 22 bp tandem repeats, respectively. Using the newly developed InDel markers with five pairs of CMS-S and their near isogenic maintainer line, we were able to easily identify their respective cytotypes. The InDel markers were further examined and applied to 1,104 plants from six Korean Sorghum cultivars to identify variant cytotypes. Additionally, the phylogenetic analysis of seven Sorghum species with complete cp genome sequences, including wild species, indicated that CMS-S and CMS-N contained Milo and Kafir cytotypes that might be hybridized from S. propinquum and S. sudanese, respectively. This study can facilitate F1 hybrid cultivar development by providing breeders with reliable tools for marker-assisted selection to breed desirable Sorghum varieties. [ABSTRACT FROM AUTHOR]

Pollen development and its fertility are obligatory conditions for the reproductive success of flowing plants. Sucrose transporter 3 (OsSUT3) is known to be preferentially expressed and may play critical role in developing pollen. A 31-bp InDel was identified as a unique variation and was shown to be responsible for the expression of downstream gene in our previous study. In this study, to analyze the changes of gene expression triggered by 31-bp InDel during pollen development, two vectors (p385-In/Del::OsSUT3-GUS) were constructed and then stably introduced into rice. Histochemical and quantitative real-time PCR (qRT-PCR) analysis of transgenic plants showed that 31-bp deletion drastically reduced the expressions of downstream genes, including both OsSUT3 and GUS in rice panicle at booting stage, especially that of OsSUT3. The transcriptome profile of two types of panicles at booting stage revealed a total of 1028 differentially expressed genes (DEGs) between 31-bp In and 31-bp Del transgenic plants. Further analyses showed that 397 of these genes were significantly enriched for the 'metabolic process' and 'binding'. Among them, nineteen genes had a strong relationship with starch and sucrose metabolism and were identified as candidate genes potentially associated with the starch accumulation in rice pollen, which that was also verified via qRT-PCR. In summary, 31-bp InDel plays a crucial role not only in the regulation of downstream genes but in the expression of sucrose-starch metabolizing genes in multiple biological pathways, and provides a different regulation mechanism for sucrose metabolism in pollen. [ABSTRACT FROM AUTHOR]

The double-petal varieties of ornamental pomegranate have higher ornamental value and garden development potential than the single-petal varieties but there has been no study on the genomic variation between them. This study aimed to determine the genomic variation between the two kinds of varieties and the relationship between the variation and phenotype by identifying the DNA variation of three single-petal varieties and three double-petal varieties using re-sequencing technology. The results showed that the variation number of each variety was in the order of single nucleotide polymorphisms (SNPs) > insertions and deletions (InDels) > structural variations (SVs) > copy number variations (CNVs). The number of SNPs and InDels in the double-petal varieties was significantly higher than that in the single-petal varieties, and there was no significant difference in the number of SVs and CNVs. The number of non-synonymous SNPs in the coding region (Nonsyn_CDS_SNPs) and InDels with a 3X length in the coding region (3X_shiftMutation_CDS_InDel) was significantly higher in the double-petal varieties than that in the single-petal varieties. The number of the two variants was strongly positively correlated with each morphological index that was related to the phenotypic difference between the two varieties. Nonsyn_CDS_SNPs and 3X_shiftMutation_CDS_InDel were enriched in the cell membrane system, cell periphery, and signal transduction, from which 15 candidate genes were screened. Our results provide genomic data for the study of the formation mechanism of the double-petal flower and lay a theoretical foundation for new variety breeding of ornamental pomegranate. [ABSTRACT FROM AUTHOR]

Fusarium wilt is a major devastating fungal disease of tomato (Solanum lycopersicum L.) caused by Fusarium oxysporum f. sp. lycopersici (Fol) which reduces the yield and production. Xylem sap protein 10 (XSP10) and Salicylic acid methyl transferase (SlSAMT) are two putative negative regulatory genes associated with Fusarium wilt of tomato. Fusarium wilt tolerance in tomato can be developed by targeting these susceptible (S) genes. Due to its efficiency, high target specificity, and versatility, CRISPR/Cas9 has emerged as one of the most promising techniques for knocking out disease susceptibility genes in a variety of model and agricultural plants to increase tolerance/resistance to various plant diseases in recent years. Though alternative methods, like RNAi, have been attempted to knock down these two S genes in order to confer resistance in tomato against Fusarium wilt, there has been no report of employing the CRISPR/Cas9 system for this specific intent. In this study, we provide a comprehensive downstream analysis of the two S genes via CRISPR/Cas9-mediated editing of single (XSP10 and SlSAMT individually) and dual-gene (XSP10 and SlSAMT simultaneously). Prior to directly advancing on to the generation of stable lines, the editing efficacy of the sgRNA-Cas9 complex was first validated using single cell (protoplast) transformation. In the transient leaf disc assay, the dual-gene editing showed strong phenotypic tolerance to Fusarium wilt disease with INDEL mutations than single-gene editing. In stable genetic transformation of tomato at the GE1 generation, dual-gene CRISPR transformants of XSP10 and SlSAMT primarily exhibited INDEL mutations than single-gene-edited lines. The dual-gene CRISPR-edited lines (CRELs) of XSP10 and SlSAMT at GE1 generation conferred a strong phenotypic tolerance to Fusarium wilt disease compared to single-gene-edited lines. Taken together, the reverse genetic studies in transient and stable lines of tomato revealed that, XSP10 and SlSAMT function together as negative regulators in conferring genetic tolerance to Fusarium wilt disease. [ABSTRACT FROM AUTHOR]

Simple Summary: The red-crowned crane is an endangered bird species in the Far East Eurasian continent and Hokkaido, Japan. Individual identification has been achieved with banding and installed transmitters in only a few cranes, though individual identification is essential for various assessments of cranes on their behavioral characteristics, for example. An insertion/deletion (InDel) mutation is a mutation ranging from 1 to 50 bp and is very useful for genetic studies. If primer sets across InDels (>20 bp) can be designed, an InDel polymorphism can be determined with conventional agarose gel electrophoresis, as reported in some plant species. We found a sufficient number of InDel primer sets to be used for individual identification and analysis of population genetics of red-crowned cranes in Hokkaido. The method has additional advantages, such as convenience and low cost, without sequencing and an expensive apparatus. Red-crowned crane Grus japonensis is an endangered species in two separate populations: the mainland population in the Eurasian continent and the island population in eastern Hokkaido, Japan. We found 11 insertion/deletion (InDel) markers in the genome of the red-crowned crane and designed primer sets across these InDels that can be analyzed with conventional agarose gel electrophoresis. Sixty-six samples of whole blood and skeletal muscle obtained from red-crowned cranes, including 12 families in eastern Hokkaido from 1994 to 2021, showed different patterns in gel images of 11 InDel PCR reactions except for two pairs. The combined non-exclusion probability of the 11 markers indicates that individuals can be determined with a probability of 99.9%. In 39 non-relative chicks, the expected heterozygosity (He) was 0.316, suggesting low genetic diversity. This might not be caused by high levels of inbreeding since the average FIS was not significantly different from zero (0.095, p = 0.075). The results suggest that the 11 InDel primer sets can be used for fairly accurate individual identification as well as genetic population analyses in red-crowned cranes in the island population. [ABSTRACT FROM AUTHOR]

Sorghum has high salt tolerance, and the localization and discovery of salt tolerance regulatory genes in sorghum is an effective way to develop and utilize saline soils. The aim of this study was to locate sorghum salt tolerance genes using a F2 segregating population combined with bulked segregation analysis (BSA) sequencing technology to explore the salt tolerance potential of sorghum. The results showed that the effective data percentage of all four samples (salt‐tolerant parent, salt‐tolerant offspring pool, salt‐sensitive parent and salt‐sensitive offspring pool) was above 99.90%. By comparing sequencing results of each of the two offspring pools with the reference genome, we found 903,025 SNP variants and 438,453 InDel variants between the two offspring pools. A total of 163 polymorphic loci were found in the genome‐wide screening of SNP‐index values close to 0.75 for the salt‐tolerant offspring pool and 0.25 for the non‐salt‐tolerant offspring pool. A total of 20 candidate genes were screened according to ANNOVAR annotation results. The Sobic.001G283700 gene encoding glycerol‐3‐phosphate acyltransferases (GPATs) was found to be closely associated with salt stress tolerance. [ABSTRACT FROM AUTHOR]

Dynein Axonemal Heavy Chain 1 (DNAH1) encodes proteins which provide structural support for the physiological function and motor structure of spermatozoa (hereafter referred to as sperm) and ova. This study found that three single nucleotide polymorphisms (SNPs), the 27-bp insertion/deletion (InDel) mutations and three exonic copy number variations (CNVs) within DNAH1 were significantly associated with litter size of Shaanbei white cashmere goats (n = 1101). Goats with the wildtypes of these three SNPs had higher litter sizes than other carriers (p < 0.05). II genotype of the 27-bp InDel had the highest litter size compared with ID carriers (p = 0.000022). The gain genotype had the largest litter sizes compared with the loss or medium carriers for the three CNV mutations (p < 0.01). Individuals with the AA-TT-CC-II-M1-M2-M3 and AA-TT-CC-II-G1-G2-M3 combination genotypes had larger litter sizes compared with the other genotypes. This study also showed the DNAH1 expression in mothers of multiple kids was higher than mothers of single kids. These three SNPs, the 27-bp InDel and three CNVs in DNAH1 could be used as molecular markers for the selection of goat reproductive traits. [ABSTRACT FROM AUTHOR]

Prime editor (PE), a versatile editor that allows the insertion and deletion of arbitrary sequences, and all 12-point mutations without double-strand breaks (DSB) and a donor template, dramatically enhances research capabilities. PE combines nickase Cas9(H840A) and reverse transcriptase (RT), along with prime editing guide RNA (pegRNA). It has been reported in several plant species, but a weak editing efficiency has led to a decrease in applications. This study reports an optimized-prime editor (O-PE) for endogenous gene editing in Arabidopsis thaliana cells, with an average 1.15% editing efficiency, which is 16.4-fold higher than previously reported. Meanwhile, we observed an increase in indels when testing alternative reverse transcriptase and found out that nCas9(H840A) fused to non-functional reverse transcriptase was responsible for the increase. This work develops an efficient prime editor for plant cells and provides a blueprint for applying PE in other photoautotrophic cells, such as microalgae, that have a high industrial value. [ABSTRACT FROM AUTHOR]

Salt-alkali tolerance is one of the important breeding traits of Portunus trituberculatus. Identification of molecular markers linked to salt-alkali tolerance is prerequisite to develop such molecular marker-assisted breeding. In this study, Bulked Segregant Analysis (BSA) was used to screen molecular markers associated with salt-alkali tolerance trait in P. trituberculatus. Two DNA mixing pools with significant difference in salt-alkali tolerance were prepared and 94.83G of high-quality sequencing data was obtained. 855 SNPs and 1051 Indels were firstly selected as candidate markers by BSA analysis, out of which, 20 markers were further selected via △index value (close to 0 or 1) and eight of those were successfully verified. In addition, based on the located information of the markers in genome, eight candidate genes related to salt-alkali tolerance were anchored including ubiquitin-conjugating enzyme, aspartate–tRNA ligase, vesicle-trafficking protein, and so on. qPCR results showed that the expression patterns of all these genes changed significantly after salt-alkali stress, suggesting that they play certain roles in salt-alkali adaptation. Our results will provide applicable markers for molecular marker-assisted breeding and help to clarify the mechanisms of salt-alkali adaptation of P. trituberculatus. [ABSTRACT FROM AUTHOR]

Accumulation of somatic hypermutation (SHM) is the primary mechanism to enhance the binding affinity of antibodies to antigens in vivo. However, the structural basis of the effects of many SHMs remains elusive. Here, we integrated atomistic molecular dynamics (MD) simulation and data mining to build a high-throughput structural bioinformatics pipeline to study the effects of individual and combination SHMs on antibody conformation, flexibility, stability, and affinity. By applying this pipeline, we characterized a common mechanism of modulation of heavy-light pairing orientation by frequent SHMs at framework positions 39H, 91H, 38L, and 87L through disruption of a conserved hydrogen-bond network. Q39LH alone and in combination with light chain framework 4 (FWR4L) insertions further modulated the elbow angle between variable and constant domains of many antibodies, resulting in improved binding affinity for a subset of anti-HIV-1 antibodies. Q39LH also alleviated aggregation induced by FWR4L insertion, suggesting remote epistasis between these SHMs. Altogether, this study provides tools and insights for understanding antibody affinity maturation and for engineering functionally improved antibodies. [ABSTRACT FROM AUTHOR]

Background In diploid organisms, whole-genome haplotype assembly relies on the accurate identification and assignment of heterozygous single-nucleotide polymorphism alleles to the correct homologous chromosomes. This appropriate phasing of these alleles ensures that combinations of single-nucleotide polymorphisms on any chromosome, called haplotypes, can then be used in downstream genetic analysis approaches including determining their potential association with important phenotypic traits. A number of statistical algorithms and complementary computational software tools have been developed for whole-genome haplotype construction from genomic sequence data. However, many algorithms lack the ability to phase long haplotype blocks and simultaneously achieve a competitive accuracy. Results In this research we present HaploMaker, a novel reference-based haplotype assembly algorithm capable of accurately and efficiently phasing long haplotypes using paired-end short reads and longer Pacific Biosciences reads from diploid genomic sequences. To achieve this we frame the problem as a directed acyclic graph with edges weighted on read evidence and use efficient path traversal and minimization techniques to optimally phase haplotypes. We compared the HaploMaker algorithm with 3 other common reference-based haplotype assembly tools using public haplotype data of human individuals from the Platinum Genome project. With short-read sequences, the HaploMaker algorithm maintained a competitively low switch error rate across all haplotype lengths and was superior in phasing longer genomic regions. For longer Pacific Biosciences reads, the phasing accuracy of HaploMaker remained competitive for all block lengths and generated substantially longer block lengths than the competing algorithms. Conclusions HaploMaker provides an improved haplotype assembly algorithm for diploid genomic sequences by accurately phasing longer haplotypes. The computationally efficient and portable nature of the Java implementation of the algorithm will ensure that it has maximal impact in reference-sequence–based haplotype assembly applications. [ABSTRACT FROM AUTHOR]

Simple Summary: Eriophyoid mites are microscopic herbivores associated with higher plants. Some of them are serious pests due to their ability to vector viruses and cause other damage to host plants. Mites of the genus Trisetacus are widespread parasites of conifers. They usually live in buds, cones, and rarely within needles of Pinaceae (pine family) and Cupressaceae (cypress family). We discovered a new species, Trisetacus indelisn. sp., severely damaging seeds of three North American junipers in the western USA. This species possesses two morphologically different forms of females and has two deletion mutations in the gene cytochrome oxidase subunit I (Cox1). Such mutations are rare in eriophyoids and were previously detected only in two pestiferous species from palms and hazelnut. Our molecular-phylogenetic analyses determine the closest known relatives of the new species and suggest that Old and New World Trisetacus independently transitioned to living in seeds of junipers. Additionally we show that reconstruction of the phylogeny of Eriophyoidea based on one gene, Cox1, produces a poorly-resolved but biologically consistent tree topology to hypothesize the evolution of Eriophyoidea. Overall, our study improves our understanding of the diversity of conifer-inhabiting mites and indicates further needs in investigating the phylogeny of Eriophyoidea. Eriophyoid mites of the genus Trisetacus Keifer are widespread parasites of conifers. A new oligophagous species, T. indelis n. sp., was discovered severely damaging seeds of North American junipers (Juniperus osteosperma, J. occidentalis, and J. californica) in the western USA. It has two codon deletions in the mitochondrial gene Cox1 rarely detected in Eriophyoidea and includes distinct morphological dimorphism of females. A phylogenetic analysis based on amino acid alignment of translated Cox1 sequences using a large set of out-groups (a) determined that two North American congeners, T. batonrougei and T. neoquadrisetus, were the closest known relatives of T. indelis n. sp., and (b) indicated that Old and New World seed-inhabiting Trisetacus from junipers do not form a distinct clade, suggesting a possible independent transition to living in seeds of junipers in America and Eurasia by Trisetacus spp. Our analysis produced a new topology consistent with a scenario assuming gradual reduction of prodorsal shield setation in Eriophyoidea and an ancient switch from gymnosperms to other hosts. Additionally, our analysis did not support monophyly of Trisetacus; recovered a new host-specific, moderately supported clade comprising Trisetacus and Nalepellinae (Nalepella + Setoptus) associated with Pinaceae; and questioned the monophyly of Trisetacus associated with Cupressaceae. [ABSTRACT FROM AUTHOR]