Genetic Crosses Research Articles

Myeloid autophagy genes protect mice against fatal TNF- and LPS-induced cytokine storm syndromes

ABSTRACT Macroautophagy/autophagy regulates inflammation via multiple mechanisms, including lysosomal degradation of specific cellular components. Certain autophagy gene “cassettes” also participate in non-canonical processes to mediate important biological activities. While select autophagy genes in myeloid cells have been implicated in protecting mice in models of cytokine storm syndromes (CSS), a more extensive genetic analysis of the autophagy pathway for this disorder has not been reported to date. We determined that multiple canonical autophagy genes in the myeloid compartment protected against fatal disease from both intravenous TNF and intraperitoneal LPS, with the notable exception that Atg14 was dispensable for the latter. Serum cytokine analyses and genetic crosses further revealed distinct mechanisms contribute to the hypersensitivity of autophagy gene-deficient mice in these CSS models. Surprisingly, TNF was dispensable for the increased mortality of myeloid 5-deficient mice challenged with LPS. Tissue-specific ablation of Atg5 in cells expressing ITGAX/CD11c and LYZ2/LYSM, but not S100A8/MRP8, defined a myeloid subset that protected against TNF, while protection against LPS was conferred by Atg5 in a distinct subset of LYZ2-expressing cells. Together, this study identifies autophagy gene sets and specific cell types that protect against fatal inflammation due to CSS, highlighting important differences in two commonly used murine models of the disorder. Abbreviations ATG5: autophagy related 5; ATG7: autophagy related 7; ATG14: autophagy related 14; ATG16L1: autophagy related 16-like 1 (S. cerevisiae); BECN1: beclin 1, autophagy related; CASP1: caspase 1; CASP4/CASP11: caspase 4, apoptosis-related cysteine peptidase; CIM: conditionally immortalized macrophage; CLP: cecal ligation and puncture; CSS: cytokine storm syndrome; DC: dendritic cell; IFNG/IFNγ: interferon gamma; IFNGR1: interferon gamma receptor 1; ip: intraperitoneal; iv: intravenous; IL12/p70: interleukin 12, p70 heterodimer; IL18: Interleukin 18; ITGAX/CD11c: integrin alpha X; LAP: LC3-associated phagocytosis; LPS: lipopolysaccharide; LYZ2/LYSM: lysozyme 2; MAP1LC3A/LC3: microtubule-associated protein 1 light chain 3 alpha; RB1CC1/FIP200: RB1-inducible coiled-coil 1; S100A8/MRP8: S100 calcium binding protein A8 (calgranulin A); TICAM1/TRIF: TIR domain containing adaptor molecule 1; TLR4: toll-like receptor 4; TNF: tumor necrosis factor.

The double round-robin population unravels the genetic architecture of grain size in barley.

Grain number, size and weight primarily determine the yield of barley. Although the genes regulating grain number are well studied in barley, the genetic loci and the causal gene for sink capacity are poorly understood. Therefore, the primary objective of our work was to dissect the genetic architecture of grain size and weight in barley. We used a multi-parent population developed from a genetic cross between 23 diverse barley inbreds in a double round-robin design. Seed size-related parameters such as grain length, grain width, grain area and thousand-grain weight were evaluated in the HvDRR population comprising 45 recombinant inbred line sub-populations. We found significant genotypic variation for all seed size characteristics, and observed 84% or higher heritability across four environments. The quantitative trait locus (QTL) detection results indicate that the genetic architecture of grain size is more complex than previously reported. In addition, both cultivars and landraces contributed positive alleles at grain size QTLs. Candidate genes identified using genome-wide variant calling data for all parental inbred lines indicated overlapping and potential novel regulators of grain size in cereals. Furthermore, our results indicated that sink capacity was the primary determinant of grain weight in barley.

Allele specific PCR for a major marker of levamisole resistance in Haemonchus contortus

Haemonchus contortus is a haematophagous parasitic nematode that infects small ruminants and causes significant animal health concerns and economic losses within the livestock industry on a global scale. Treatment primarily depends on broad-spectrum anthelmintics, however, resistance is established or rapidly emerging against all major drug classes. Levamisole (LEV) remains an important treatment option for parasite control, as resistance to LEV is less prevalent than to members of other major classes of anthelmintics. LEV is an acetylcholine receptor (AChR) agonist that, when bound, results in paralysis of the worm. Numerous studies implicated the AChR sub-unit, ACR-8, in LEV sensitivity and in particular, the presence of a truncated acr-8 transcript or a deletion in the acr-8 locus in some resistant isolates. Recently, a single non-synonymous SNP in acr-8 conferring a serine-to-threonine substitution (S168T) was identified that was strongly associated with LEV resistance. Here, we investigate the role of genetic variation at the acr-8 locus in a controlled genetic cross between the LEV susceptible MHco3(ISE) and LEV resistant MHco18(UGA2004) isolates of H. contortus. Using single worm PCR assays, we found that the presence of S168T was strongly associated with LEV resistance in the parental isolates and F3 progeny of the genetic cross surviving LEV treatment. We developed and optimised an allele-specific PCR assay for the detection of S168T and validated the assay using laboratory isolates and field samples that were phenotyped for LEV resistance. In the LEV-resistant field population, a high proportion (>75%) of L3 encoded the S168T variant, whereas the variant was absent in the susceptible isolates studied. These data further support the potential role of acr-8 S168T in LEV resistance, with the allele-specific PCR providing an important step towards establishing a sensitive molecular diagnostic test for LEV resistance.

Eudicot primary cell wall glucomannan is related in synthesis, structure, and function to xyloglucan.

Hemicellulose polysaccharides influence assembly and properties of the plant primary cell wall (PCW), perhaps by interacting with cellulose to affect the deposition and bundling of cellulose fibrils. However, the functional differences between plant cell wall hemicelluloses such as glucomannan, xylan, and xyloglucan (XyG) remain unclear. As the most abundant hemicellulose, XyG is considered important in eudicot PCWs, but plants devoid of XyG show relatively mild phenotypes. We report here that a patterned β-galactoglucomannan (β-GGM) is widespread in eudicot PCWs and shows remarkable similarities to XyG. The sugar linkages forming the backbone and side chains of β-GGM are analogous to those that make up XyG, and moreover, these linkages are formed by glycosyltransferases from the same CAZy families. Solid-state nuclear magnetic resonance indicated that β-GGM shows low mobility in the cell wall, consistent with interaction with cellulose. Although Arabidopsis β-GGM synthesis mutants show no obvious growth defects, genetic crosses between β-GGM and XyG mutants produce exacerbated phenotypes compared with XyG mutants. These findings demonstrate a related role of these two similar but distinct classes of hemicelluloses in PCWs. This work opens avenues to study the roles of β-GGM and XyG in PCWs.

Genetic Variation in Heat Tolerance of the Coral Platygyra Daedalea Indicates Potential for Adaptation to Ocean Warming

Ocean warming represents the greatest threat to the persistence of reef ecosystems. Most coral populations are projected to experience temperatures above their current bleaching thresholds annually by 2050. Adaptation to higher temperatures is necessary if corals are to persist in a warming future. While many aspects of heat stress have been well studied, few data are available for predicting the capacity for adaptive cross-generational responses in corals. Consistent sets of heat tolerant genomic markers that reliably predict thermal tolerance have yet to be identified. To address this knowledge gap, we quantified the heritability and genetic variation associated with heat tolerance in Platygyra daedalea from the Great Barrier Reef. We tracked the survival of ten quantitative genetic crosses of larvae produced form six parental colonies in a heat tolerance selection experiment. We also identified allelic shifts in heat-selected (35°C) survivors compared with paired, non-selected controls (27°C). The narrow-sense heritability of survival under heat stress was 0.66 and a total of 1,069 single nucleotide polymorphisms (SNPs) were associated with different survival probabilities. While 148 SNPs were shared between several experimental crosses, no common SNPs were identified for all crosses, which suggests that specific combinations of many markers are responsible for heat tolerance. However, we found two regions that overlap with previously identified loci associated with heat tolerance in Persian Gulf populations of P. daedalea, which reinforces the importance of these markers for heat tolerance. These results illustrate the importance of high heritability and the complexity of the genomic architecture underpinning host heat tolerance. These findings suggest that this P. daedalea population has the genetic prerequisites for adaptation to increasing temperatures. This study also provides knowledge for the development of high throughput genomic tools which may screen for variation within and across populations to enhance adaptation through assisted gene flow and assisted migration.

Keystone Malaria Symposium 2022: a vibrant discussion of progress made and challenges ahead from drug discovery to treatment

Keystone Malaria Symposium 2022: a vibrant discussion of progress made and challenges ahead from drug discovery to treatment

A Multitrait Locus Regulates Sarbecovirus Pathogenesis.

ABSTRACTInfectious diseases have shaped the human population genetic structure, and genetic variation influences the susceptibility to many viral diseases. However, a variety of challenges have made the implementation of traditional human Genome-wide Association Studies (GWAS) approaches to study these infectious outcomes challenging. In contrast, mouse models of infectious diseases provide an experimental control and precision, which facilitates analyses and mechanistic studies of the role of genetic variation on infection. Here we use a genetic mapping cross between two distinct Collaborative Cross mouse strains with respect to severe acute respiratory syndrome coronavirus (SARS-CoV) disease outcomes. We find several loci control differential disease outcome for a variety of traits in the context of SARS-CoV infection. Importantly, we identify a locus on mouse chromosome 9 that shows conserved synteny with a human GWAS locus for SARS-CoV-2 severe disease. We follow-up and confirm a role for this locus, and identify two candidate genes, CCR9 and CXCR6, that both play a key role in regulating the severity of SARS-CoV, SARS-CoV-2, and a distantly related bat sarbecovirus disease outcomes. As such we provide a template for using experimental mouse crosses to identify and characterize multitrait loci that regulate pathogenic infectious outcomes across species.

Molecular Advancements Establishing Chlamydomonas as a Host for Biotechnological Exploitation

Chlamydomonas reinhardtii is emerging as a production platform for biotechnological purposes thanks to recent achievements, which we briefly summarize in this review. Firstly, robust nuclear transgene expression is now possible because several impressive improvements have been made in recent years. Strains allowing efficient and stable nuclear transgene expression are available and were recently made more amenable to rational biotechnological approaches by enabling genetic crosses and identifying their causative mutation. The MoClo synthetic biology strategy, based on Golden Gate cloning, was developed for Chlamydomonas and includes a growing toolkit of more than 100 genetic parts that can be robustly and rapidly assembled in a predefined order. This allows for rapid iterative cycles of transgene design, building, testing, and learning. Another major advancement came from various findings improving transgene design and expression such as the systematic addition of introns into codon-optimized coding sequences. Lastly, the CRISPR/Cas9 technology for genome editing has undergone several improvements since its first successful report in 2016, which opens the possibility of optimizing biosynthetic pathways by switching off competing ones. We provide a few examples demonstrating that all these recent developments firmly establish Chlamydomonas as a chassis for synthetic biology and allow the rewiring of its metabolism to new capabilities.

Rapid and specific degradation of endogenous proteins in mouse models using auxin-inducible degrons.

Auxin-inducible degrons are a chemical genetic tool for targeted protein degradation and are widely used to study protein function in cultured mammalian cells. Here, we develop CRISPR-engineered mouse lines that enable rapid and highly specific degradation of tagged endogenous proteins in vivo. Most but not all cell types are competent for degradation. By combining ligand titrations with genetic crosses to generate animals with different allelic combinations, we show that degradation kinetics depend upon the dose of the tagged protein, ligand, and the E3 ligase substrate receptor TIR1. Rapid degradation of condensin I and II - two essential regulators of mitotic chromosome structure - revealed that both complexes are individually required for cell division in precursor lymphocytes, but not in their differentiated peripheral lymphocyte derivatives. This generalisable approach provides unprecedented temporal control over the dose of endogenous proteins in mouse models, with implications for studying essential biological pathways and modelling drug activity in mammalian tissues.

Homologous chromosome associations in domains before meiosis could facilitate chromosome recognition and pairing in wheat

The increasing human population demands an increase in crop yields that must be implemented through breeding programmes to ensure a more efficient and sustainable production of agro-food products. In the framework of breeding, genetic crosses are developed between cultivated species such as wheat and their relative species that are used as genetic donors to transfer desirable agronomic traits into the crop. Unfortunately, interspecific associations between chromosomes from the donor species and the cultivar are rare during meiosis, the process to produce gametes in organisms with sexual reproduction, hampering the transfer of genetic variability into wheat. In addition, little is known about how homologous (equivalent) chromosomes initiate interaction and recognition within the cell nucleus to enter meiosis. In this context, we aim to get insight into wheat chromatin structure, particularly the distribution of homologous chromosomes within the cell nucleus and their putative interactions in premeiotic stages to facilitate chromosome associations and recombination at the beginning of meiosis. Cytogenetics allows the study of both the structure and the behaviour of chromosomes during meiosis and is key in plant breeding. In this study we visualized an extra pair of barley homologous chromosomes in a wheat genetic background to study the spatial distribution, arrangements and interactions occurring exclusively between this pair of homologous chromosomes during premeiosis using fluorescence in situ hybridization (FISH). Our results suggest that homologous chromosomes can initiate interactions in premeiotic stages that could facilitate the processes of specific chromosome recognition and association occurring at the onset of meiosis.

Transcriptomic analyses implicate neuronal plasticity and chloride homeostasis in ivermectin resistance and response to treatment in a parasitic nematode.

The antiparasitic drug ivermectin plays an essential role in human and animal health globally. However, ivermectin resistance is widespread in veterinary helminths and there are growing concerns of sub-optimal responses to treatment in related helminths of humans. Despite decades of research, the genetic mechanisms underlying ivermectin resistance are poorly understood in parasitic helminths. This reflects significant uncertainty regarding the mode of action of ivermectin in parasitic helminths, and the genetic complexity of these organisms; parasitic helminths have large, rapidly evolving genomes and differences in evolutionary history and genetic background can confound comparisons between resistant and susceptible populations. We undertook a controlled genetic cross of a multi-drug resistant and a susceptible reference isolate of Haemonchus contortus, an economically important gastrointestinal nematode of sheep, and ivermectin-selected the F2 population for comparison with an untreated F2 control. RNA-seq analyses of male and female adults of all populations identified high transcriptomic differentiation between parental isolates, which was significantly reduced in the F2, allowing differences associated specifically with ivermectin resistance to be identified. In all resistant populations, there was constitutive upregulation of a single gene, HCON_00155390:cky-1, a putative pharyngeal-expressed transcription factor, in a narrow locus on chromosome V previously shown to be under ivermectin selection. In addition, we detected sex-specific differences in gene expression between resistant and susceptible populations, including constitutive upregulation of a P-glycoprotein, HCON_00162780:pgp-11, in resistant males only. After ivermectin selection, we identified differential expression of genes with roles in neuronal function and chloride homeostasis, which is consistent with an adaptive response to ivermectin-induced hyperpolarisation of neuromuscular cells. Overall, we show the utility of a genetic cross to identify differences in gene expression that are specific to ivermectin selection and provide a framework to better understand ivermectin resistance and response to treatment in parasitic helminths.

Somatic Genetics Analysis of Sleep in Adult Mice.

Classical forward and reverse mouse genetics require germline mutations and, thus, are unwieldy to study sleep functions of essential genes or redundant pathways. It is also time-consuming to conduct EEG/EMG-based mouse sleep screening because of labor-intensive surgeries and genetic crosses. Here, we describe a highly accurate SleepV (video) system and adeno-associated virus (AAV)-based adult brain chimeric (ABC)-expression/KO platform for somatic genetics analysis of sleep in adult male or female mice. A pilot ABC screen identifies CREB and CRTC1, of which constitutive or inducible expression significantly reduces quantity and/or quality of non-rapid eye movement sleep. Whereas ABC-KO of exon 13 of Sik3 by AAV-Cre injection in Sik3-E13flox/flox adult mice phenocopies Sleepy (Sik3Slp/+) mice, ABC-CRISPR of Slp/Sik3 reverses hypersomnia of Sleepy mice, indicating a direct role of SLP/SIK3 kinase in sleep regulation. Multiplex ABC-CRISPR of both orexin/hypocretin receptors causes narcolepsy episodes, enabling one-step analysis of redundant genes in adult mice. Therefore, this somatic genetics approach should facilitate high-throughput analysis of sleep regulatory genes, especially for essential or redundant genes, in adult mice by skipping mouse development and minimizing genetic crosses.SIGNIFICANCE STATEMENT The molecular mechanisms of mammalian sleep regulation remain unclear. Classical germline mouse genetics are unwieldy to study sleep functions of essential genes or redundant pathways. The EEG/EMG-based mouse sleep screening is time-consuming because of labor-intensive surgeries and lengthy genetic crosses. To overcome these “bottlenecks,” we developed a highly accurate video-based sleep analysis system and adeno-associated virus-mediated ABC-expression/KO platform for somatic genetics analysis of sleep in adult mice. These methodologies facilitate rapid identification of sleep regulatory genes, but also efficient mechanistic studies of the molecular pathways of sleep regulation in mice.

Enabling Medicago truncatula forward genetics: identification of genetic crossing partner for R108 and development of mapping resources for Tnt1 mutants.

Though Medicago truncatula Tnt1 mutants are widely used by researchers in the legume community, they are mainly used for reverse genetics because of the availability of the BLAST-searchable large-scale flanking sequence tags database. However, these mutants should have also been used extensively for forward genetic screens, an effort that has been hindered due to the lack of a compatible genetic crossing partner for the M. truncatula genotype R108, from which Tnt1 mutants were generated. In this study, we selected three Medicago HapMap lines (HM017, HM018 and HM022) and performed reciprocal genetic crosses with R108. After phenotypic analyses in F1 and F2 progenies, HM017 was identified as a compatible crossing partner with R108. By comparing the assembled genomic sequences of HM017 and R108, we developed and confirmed 318 Indel markers evenly distributed across the eight chromosomes of the M. truncatula genome. To validate the effectiveness of these markers, by employing the map-based cloning approach, we cloned the causative gene in the dwarf mutant crs isolated from the Tnt1 mutant population, identifying it as gibberellin 3-β-dioxygenase 1, using some of the confirmed Indel markers. The primer sequences and the size difference of each marker were made available for users in the web-based database. The identification of the crossing partner for R108 and the generation of Indel markers will enhance the forward genetics and the overall usage of the Tnt1 mutants.

A Malaria Parasite Cross Reveals Genetic Determinants of Plasmodium falciparum Growth in Different Culture Media.

What genes determine in vitro growth and nutrient utilization in asexual blood-stage malaria parasites? Competition experiments between NF54, clone 3D7, a lab-adapted African parasite, and a recently isolated Asian parasite (NHP4026) reveal contrasting outcomes in different media: 3D7 outcompetes NHP4026 in media containing human serum, while NHP4026 outcompetes 3D7 in media containing AlbuMAX, a commercial lipid-rich bovine serum formulation. To determine the basis for this polymorphism, we conducted parasite genetic crosses using humanized mice and compared genome-wide allele frequency changes in three independent progeny populations cultured in media containing human serum or AlbuMAX. This bulk segregant analysis detected three quantitative trait loci (QTL) regions [on chromosome (chr) 2 containing aspartate transaminase AST; chr 13 containing EBA-140; and chr 14 containing cysteine protease ATG4] linked with differential growth in serum or AlbuMAX in each of the three independent progeny pools. Selection driving differential growth was strong (s = 0.10 – 0.23 per 48-hour lifecycle). We conducted validation experiments for the strongest QTL on chr 13: competition experiments between ΔEBA-140 and 3D7 wildtype parasites showed fitness reversals in the two medium types as seen in the parental parasites, validating this locus as the causative gene. These results (i) demonstrate the effectiveness of bulk segregant analysis for dissecting fitness traits in P. falciparum genetic crosses, and (ii) reveal intimate links between red blood cell invasion and nutrient composition of growth media. Use of parasite crosses combined with bulk segregant analysis will allow systematic dissection of key nutrient acquisition/metabolism and red blood cell invasion pathways in P. falciparum.

The Kelch-F-box protein SMALL AND GLOSSY LEAVES 1 (SAGL1) negatively influences salicylic acid biosynthesis in Arabidopsis thaliana by promoting the turn-over of transcription factor SYSTEMIC ACQUIRED RESISTANCE DEFICIENT 1 (SARD1).

Salicylic acid (SA) is a key phytohormone regulating plant immunity. Although the transcriptional regulation of SA biosynthesis has been well-studied, its post-translational regulation is largely unknown. We report that a Kelch repeats-containing F-box (KFB) protein, SMALL AND GLOSSY LEAVES 1 (SAGL1), negatively influences SA biosynthesis in Arabidopsis thaliana by mediating the proteolytic turnover of SYSTEMIC ACQUIRED RESISTANCE DEFICIENT 1 (SARD1), a master transcription factor that directly drives SA biosynthesis during immunity. Loss of SAGL1 resulted in characteristic growth inhibition. Combining metabolomic, transcriptional and phenotypic analyses, we found that SAGL1 represses SA biosynthesis and SA-mediated immune activation. Genetic crosses to mutants that are deficient in SA biosynthesis blocked the SA overaccumulation in sagl1 and rescued its growth. Biochemical and proteomic analysis identified that SAGL1 interacts with SARD1 and promotes the degradation of SARD1 in a proteasome-dependent manner. These results unravelled a critical role of KFB protein SAGL1 in maintaining SA homeostasis via controlling SARD1 stability.

Putative Condition-Dependent Viability Selection in Wild-Type Stocks of Drosophila pseudoobscura

Meiotic recombination rates vary in response to intrinsic and extrinsic factors. Recently, heat stress has been shown to reveal plasticity in recombination rates in Drosophila pseudoobscura. Here, a combination of molecular genotyping and X-linked recessive phenotypic markers were used to investigate differences in recombination rates due to heat stress. In addition, haplotypes from the genetic crosses were compared to test if they deviated from equal proportions, which would indicate viability selection. To avoid this potential bias, SNP genotyping markers overlapping the regions assayed with mutant markers were used to further investigate recombination rate. Interestingly, skews in haplotype frequency were consistent with the fixation of alleles in the wild-type stocks used that are unfit at high temperature. Evidence of viability selection due to heat stress in the wild-type haplotypes was most apparent on days 7–9 when more mutant non-crossover haplotypes were recovered in comparison to wild type (p < 0.0001). Recombination analysis using SNP markers showed days 9–10 as significantly different due to heat stress in 2 pairs of consecutive SNP markers (p = 0.018; p = 0.015), suggesting that during this time period the recombination rate is most sensitive to heat stress. This peak timing for recombination plasticity is consistent with Drosophila melanogaster based on a comparison of similarly timed key meiotic events, enabling future mechanistic work of temperature stress on recombination rate.

The transcription factor Atf1 lowers the transition barrier for nucleosome-mediated establishment of heterochromatin.

Transcription factors can exert opposite effects depending on the chromosomal context. The fission yeast transcription factor Atf1 both activates numerous genes in response to stresses and mediates heterochromatic gene silencing in the mating-type region. Investigating this context dependency, we report here that the establishment of silent heterochromatin in the mating-type region occurs at a reduced rate in the absence of Atf1 binding. Quantitative modeling accounts for the observed establishment profiles by a combinatorial recruitment of histone-modifying enzymes: locally by Atf1 at two binding sites and over the whole region by dynamically appearing heterochromatic nucleosomes, a source of which is the RNAi-dependent cenH element. In the absence of Atf1 binding, the synergy is lost, resulting in a slow rate of heterochromatin formation. The system shows how DNA-binding proteins can influence local nucleosome states and thereby potentiate long-range positive feedback on histone-modification reactions to enable heterochromatin formation over large regions in a context-dependent manner.

NanoDam identifies Homeobrain (ARX) and Scarecrow (NKX2.1) as conserved temporal factors in the Drosophila central brain and visual system.

Temporal patterning of neural progenitors is an evolutionarily conserved strategy for generating neuronal diversity. Type II neural stem cells in the Drosophila central brain produce transit-amplifying intermediate neural progenitors (INPs) that exhibit temporal patterning. However, the known temporal factors cannot account for the neuronal diversity in the adult brain. To search for missing factors, we developed NanoDam, which enables rapid genome-wide profiling of endogenously tagged proteins invivo with a single genetic cross. Mapping the targets of known temporal transcription factors with NanoDam revealed that Homeobrain and Scarecrow (ARX and NKX2.1 orthologs) are also temporal factors. We show that Homeobrain and Scarecrow define middle-aged and late INP temporal windows and play a role in cellular longevity. Strikingly, Homeobrain and Scarecrow have conserved functions as temporal factors in the developing visual system. NanoDam enables rapid cell-type-specific genome-wide profiling with temporal resolution and is easily adapted for use in higher organisms.

Admixture of divergent genomes facilitates hybridization across species in the family Brassicaceae.

Summary Hybridization and polyploidization are pivotal to plant evolution. Genetic crosses between distantly related species are rare in nature due to reproductive barriers but how such hurdles can be overcome is largely unknown. Here we report the hybrid genome structure of xBrassicoraphanus, a synthetic allotetraploid of Brassica rapa and Raphanus sativus.We performed cytogenetic analysis and de novo genome assembly to examine chromosome behaviors and genome integrity in the hybrid. Transcriptome analysis was conducted to investigate expression of duplicated genes in conjunction with epigenome analysis to address whether genome admixture entails epigenetic reconfiguration.Allotetraploid xBrassicoraphanus retains both parental chromosomes without genome rearrangement. Meiotic synapsis formation and chromosome exchange are avoided between nonhomologous progenitor chromosomes. Reconfiguration of transcription network occurs, and less divergent cis‐elements of duplicated genes are associated with convergent expression. Genome‐wide DNA methylation asymmetry between progenitors is largely maintained but, notably, B. rapa‐originated transposable elements are transcriptionally silenced in xBrassicoraphanus through gain of DNA methylation.Our results demonstrate that hybrid genome stabilization and transcription compatibility necessitate epigenome landscape adjustment and rewiring of cis–trans interactions. Overall, this study suggests that a certain extent of genome divergence facilitates hybridization across species, which may explain the great diversification and expansion of angiosperms during evolution.

Principal Component Analysis of Different Economic Traits in Layer Chicken

Background: Principal component analysis is a multivariate technique that transforms a number of possibly correlated variables into smaller number of uncorrelated variables leads to dimension reduction. Various economic traits of layer chicken are used for selection of parent birds which need to adjust for selection strategies to augment the genetic improvement. Methods: The data was collected from 2020-2021 which includes weekly body weight (g) from 0 day to 20th week and 40th week, Body weight (g) at first egg production, age at sexual maturity (days), weight of first egg (g), egg numbers at 40th week, egg weight at 40th week (g), egg numbers at 52nd week, egg weight at 52nd week (g). The least squares mean was estimated considering three different genetic groups of layer chicken (N=450, 150 each group). The main focus of this study was to identify the principal components for economic traits in layer chicken. Further varimax rotation method was applied for the transformation of components to approximate simple structure. Result: The genetic group Desi cross 1 performed better than Desi cross 2 followed by Punjab Red layer chicken. A total of three principal components were obtained which explained a total variance of 75.524%. Principal component 1 had high loads on body weight 10th week to 20th week (BW 10-BW 20) and BWSM and had a variance of 38.892%. Similarly, PC2 and PC3 explained variance of 27.072% and 9.560% respectively and had high loads on 1 week body weight to 9-week body weight (BW1-BW9) and age at sexual maturity, 40-week egg production, 52-week egg production respectively. From this study it was included that PCA can be used for selecting the economic traits for breeding purpose of layer chicken.