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Large-scale functional genomic analysis of sporulation and meiosis in Saccharomyces cerevisiae.

We have used a single-gene deletion mutant bank to identify the genes required for meiosis and sporulation among 4323 nonessential Saccharomyces cerevisiae annotated open reading frames (ORFs). Three hundred thirty-four sporulation-essential genes were identified, including 78 novel ORFs and 115 known genes without previously described sporulation defects in the comprehensive Saccharomyces Genome (SGD) or Yeast Proteome (YPD) phenotype databases. We have further divided the uncharacterized sporulation-essential genes into early, middle, and late stages of meiosis according to their requirement for IME1 induction and nuclear division. We believe this represents a nearly complete identification of the genes uniquely required for this complex cellular pathway. The set of genes identified in this phenotypic screen shows only limited overlap with those identified by expression-based studies.

Genes, Fungal↗

Clinicopathologic study of PRAD1/cyclin D1 overexpressing lymphoma with special reference to mantle cell lymphoma. A distinct molecular pathologic entity.

Mantle cell lymphomas (MCLs) are frequently associated with the overexpression of PRAD1/cyclin D1, activated by 11q13 translocation and its molecular counterpart BCL-1 gene rearrangement. We recently described the correlation of positive nuclear staining using monoclonal antibody against a PRAD1/cyclin D1 product with mRNA overexpression in MCLs. In the present study, we immunohistochemically investigated the PRAD1/cyclin D1 protein in a large series of 334 lymphoproliferative disorders, including 39 cases of MCLs on paraffin sections. Based on the cyclin D1 positivity, CD5 expression, and the morphologic features of the tumor tissue, four groups of MCL-related lesions were identified among the B-cell lymphomas examined: 36 cases with cyclin D1 overexpression, 35 (95%) of which exhibited CD5-positivity and MCL-morphology (Group 1); four cases of lymphomas with MCL morphology and CD5 expression but lacking cyclin D1 overexpression (Group II); four cases of lymphomas without cyclin D1 overexpression and surface CD5 but that fall within the morphologic boundaries of MCLs (Group III); and 11 cases of CD5-positive diffuse large cell lymphomas without cyclin D1 overexpression (Group IV). The Group I cases demonstrated quite homogeneous clinicopathologic features identical to those of MCLs. This group showed a poor prognosis (11% had 5-year survival), which is highly contrasted with that of Group II (100%). Although the four groups of MCL-related lesions sometimes overlapped in their histologic or phenotypic spectrums, each appeared to show distinct clinicopathologic and prognostic profiles. Our study provides a basis for further clarification of the nature of the neoplasms of Groups II, III, and IV. Moreover, this comprehensive study may indicate that the overexpression of PRAD1/cyclin D1 is biologically essential to defining MCLs.

Aged↗

The DND1-NANOS3 complex shapes the primordial germ cell transcriptome via a heptanucleotide sequence in mRNA 3' UTRs.

The RNA-binding proteins DND1 and NANOS3 are essential for primordial germ cell survival1-5. Their co-immunoprecipitation and overlapping loss-of-function phenotypes suggest joint function6-8, yet how they co-regulate target mRNAs remains unclear. Here, we developed Tandem PAR-CLIP and identified a DND1-NANOS3 ribonucleoprotein that specifically recognizes an AUGAAUU heptanucleotide on target mRNAs, termed the NANOS3-dependent DND1 Recognition Element (N3-DRE). mRNAs containing 3'-UTR N3-DREs are aberrantly upregulated in DND1- or NANOS3-deficient germ cells and encode key cell-cycle and epigenome regulators, such as CDK1. Genome editing showed that the N3-DRE is essential for Cdk1 repression in mouse PGCs in vivo. A 1.7-Å crystal structure of the ternary complex of DND1, NANOS3, and CDK1-N3-DRE RNA revealed a continuous RNA-binding surface that confers high-affinity, sequence-specific recognition. Together, these findings define the molecular and functional basis of N3-DRE-mediated mRNA regulation in germ cell development. Moreover, we provide a paradigm of two RNA-binding proteins with low (DND1) or no (NANOS3) intrinsic sequence-specificity, jointly building a high-information-content RNA sequence motif that is different from the sum of their individual preferences. Because RNA-binding protein specificities are typically studied individually9-13, rather than in the context of ribonucleoproteins, this type of "two-factor authorization" may be an underappreciated mechanism to protect posttranscriptional gene regulatory networks from aberrant expression of an individual ribonucleoprotein component.

Journal Article↗

Hybridization between subspecies of waterbuck (Kobus ellipsiprymnus) in zones of overlap with limited introgression.

Two subspecies of waterbuck (Kobus ellipsiprymnus), common (Kobus ellipsiprymnus ellipsiprymnus) and defassa (Kobus ellipsiprymnus defassa), are recognized based on differences in rump pattern, coat colour and geographical distribution. These forms are parapatrically distributed with an area of range overlap in East Africa, where phenotypically intermediate populations occur. Variation in 478 bp of the mitochondrial DNA control region and 14 polymorphic microsatellite loci were used to describe the genetic structure and phylogeographical pattern of the species, and to assess if the intermediate populations are the results of hybridization. In total, 186 individuals from 11 localities were analysed. A high degree of genetic differentiation was found between subspecies, although this was most evident from the microsatellite data. Hybridization was suggested in the phenotypically and geographically intermediate Nairobi NP population in Kenya. A neighbour-joining (NJ) tree based on microsatellite population genetic distances grouped Nairobi between the common and defassa populations, and a Bayesian analysis clearly showed introgression. Individuals sampled in Samburu NP, Kenya, had a common waterbuck phenotype, but introgression was suggested by both markers. Although a high degree of maternal defassa input was indicated from the sequence data, the Samburu population grouped with the common waterbuck in the microsatellite population genetic distance tree, with high support. Analyses of linkage disequilibrium and maximum-likelihood estimates of genetic drift suggested that admixture between subspecies is a recent event. The fact that introgression is limited between subspecies could be caused by chromosomal differences, hindering gene flow between common and defassa waterbuck.

Africa, Eastern↗

The conserved Xanthomonas campestris pv. vesicatoria effector protein XopX is a virulence factor and suppresses host defense in Nicotiana benthamiana.

Nicotiana benthamiana leaves display a visible plant cell death response when infiltrated with a high titer inoculum of the non-host pathogen, Xanthomonas campestris pv. vesicatoria (Xcv). This visual phenotype was used to identify overlapping cosmid clones from a genomic cosmid library constructed from the Xcv strain, GM98-38. Individual cosmid clones from the Xcv library were conjugated into X. campestris pv. campestris (Xcc) and exconjugants were scored for an altered visual high titer inoculation response in N. benthamiana. The molecular characterization of the cosmid clones revealed that they contained a novel gene, xopX, that encodes a 74-kDa type III secretion system (TTSS) effector protein. Agrobacterium-mediated transient expression of XopX in N. benthamiana did not elicit the plant cell death response although detectable XopX protein was produced. Interestingly, the plant cell death response occurred when the xopX Agrobacterium-mediated transient expression construct was co-inoculated with strains of either XcvDeltaxopX or Xcc, both lacking xopX. The co-inoculation complementation of the plant cell death response also depends on whether the Xanthomonas strains contain an active TTSS. Transgenic 35S-xopX-expressing N. benthamiana plants also have the visible plant cell death response when inoculated with the non-xopX-expressing strains XcvDeltaxopX and Xcc. Unexpectedly, transgenic 35S-xopX N. benthamiana plants displayed enhanced susceptibility to bacterial growth of Xcc as well as other non-xopX-expressing Xanthomonas and Pseudomonas strains. This result is also consistent with the increase in bacterial growth on wild type N. benthamiana plants observed for Xcc when XopX is expressed in trans. Furthermore, XopX contributes to the virulence of Xcv on host pepper (Capsicum annuum) and tomato (Lycopersicum esculentum) plants. We propose that the XopX bacterial effector protein targets basic innate immunity in plants, resulting in enhanced plant disease susceptibility.

Amino Acid Sequence↗

Effects of acute and repeated ethanol exposures on the locomotor activity of BXD recombinant inbred mice.

Investigations of ethanol's (EtOH's) complex response profile, including locomotor and other effects, are likely to lead to a more in-depth understanding of the constituents of alcohol addiction. Locomotor activity responses to acute and repeated EtOH (2 g/kg, ip) exposures were measured in BXD recombinant inbred (RI) mice and their C57BL/6J (B6) and DBA/2J (D2) progenitors. Both the acute response and the change in initial EtOH response with repeated treatments were strain-dependent. The coefficient of genetic determination was 0.38-0.49 for initial locomotor response to EtOH, and 0.29 for change in response. Changes in response were largely attributable to sensitization of locomotor stimulation. Quantitative trait loci (QTL) analyses identified significant marker associations with basal activity, acute locomotor response, and change in response. Markers were for QTL on several chromosomes, and there was only one case of overlap in marker associations among phenotypes. Acute locomotor response and locomotor sensitization were negatively correlated with 3% EtOH preference drinking data collected in BXD RI strains. Overall, these results demonstrate locomotor sensitization induced by EtOH, suggest independence of genetic determination of locomotor responses to acute and repeated EtOH exposure, and partially support a relationship between reduced sensitivity to the locomotor stimulant/sensitizing effects of EtOH and EtOH consumption.

Alcoholism↗

Presence of somatic hypermutation and activation-induced cytidine deaminase in acute lymphoblastic leukemia L2 with t(14;18)(q32;q21).

AIM: Acute lymphoblastic leukemia (ALL) with L2 (FAB) morphology has rarely been reported to show t(14;18)(q32;q21). We aimed to delineate the stage at which this type of ALL is derived in B-lineage differentiation. METHODS: The somatic hypermutation (SHM) of the variable region of immunoglobulin heavy chain (IgV(H)) gene and the expression of terminal deoxynucleotidyl transferase (TdT), recombination-activating gene 1 and 2 (RAG-1 and -2), and activation-induced cytidine deaminase (AID) were investigated in three cell lines and two fresh samples, including a pair of matched fresh and cell line cells. RESULTS: TdT, RAG-1, and RAG-2 were variably expressed. AID was expressed in four of five samples. SHM of the IgV(H) gene was found in all samples with high average frequency (11.84%) comparable with that in follicular lymphoma. Ongoing mutation was seen in two fresh samples. CONCLUSION: As AID and SHM are generally regarded as properties exhibited by mature B cells, the presence of AID and SHM in this study seems to be incompatible with the general understanding of the early stage derivation of ALL in B-lineage differentiation. The results here give some insight into the relationship between disease type (ALL or lymphoma) and derivation stage, the overlapping of the early stage phenotype and the mature genomic characteristics, and the probable relationship between the mechanism of the occurrence of t(14;18)(q32;q21) and the machinery causing SHM.

Base Sequence↗

Further characterization of complement regulator-acquiring surface proteins of Borrelia burgdorferi.

The three genospecies Borrelia burgdorferi, Borrelia garinii, and Borrelia afzelii, all causative agents of Lyme disease, differ in their susceptibilities to human complement-mediated lysis. We recently reported that serum resistance of borrelias correlates largely with their ability to bind the human complement regulators FHL-1/reconectin and factor H. To date, two complement regulator-acquiring-proteins (CRASP-1 and CRASP-2) have been identified in serum-resistant B. afzelii isolates (P. Kraiczy, C. Skerka, M. Kirschfink, V. Brade, and P. F. Zipfel, Eur. J. Immunol. 31:1674-1684, 2001). Here, we present a comprehensive study of the CRASPs detectable in both serum-resistant and intermediate serum-sensitive B. afzelii and B. burgdorferi isolates. These CRASPs were designated according to the genospecies either as BaCRASPs, when derived from B. afzelii, or as BbCRASPs, for proteins identified in B. burgdorferi isolates. Each borrelial isolate expresses distinct CRASPs that can be differentiated by their mobility and binding phenotypes. A detailed comparison reveals overlapping and even identical binding profiles for BaCRASP-1 (27.5 kDa), BbCRASP-1 (25.9 kDa), and BbCRASP-2 (23.2 kDa), which bind FHL-1/reconectin strongly and interact weakly with factor H. In contrast, two B. afzelii proteins (BaCRASP-4 [19.2 kDa] and BaCRASP-5 [22.5 kDa]) and three B. burgdorferi proteins (BbCRASP-3 [19.8 kDa], BbCRASP-4 [18.5 kDa], and BbCRASP-5 [17.7 kDa]) bind factor H but not FHL-1/reconectin. Most CRASPs bind both human immune regulators at their C-terminal ends. Temperature-dependent up-regulation of CRASPs (BaCRASP-1, BaCRASP-2, and BaCRASP-5) is detected in low-passage borrelias cultured at 33 or 37 degrees C compared with those cultured at 20 degrees C. The characterization of the individual CRASPs on the molecular level is expected to identify new virulence factors and potential vaccine candidates.

Bacterial Proteins↗

Mice doubly deficient for the Polycomb Group genes Mel18 and Bmi1 reveal synergy and requirement for maintenance but not initiation of Hox gene expression.

Polycomb group genes were identified as a conserved group of genes whose products are required in multimeric complexes to maintain spatially restricted expression of Hox cluster genes. Unlike in Drosophila, in mammals Polycomb group (PcG) genes are represented as highly related gene pairs, indicative of duplication during metazoan evolution. Mel18 and Bmi1 are mammalian homologs of Drosophila Posterior sex combs. Mice deficient for Mel18 or Bmi1 exhibit similar posterior transformations of the axial skeleton and display severe immune deficiency, suggesting that their gene products act on overlapping pathways/target genes. However unique phenotypes upon loss of either Mel18 or Bmi1 are also observed. We show using embryos doubly deficient for Mel18 and Bmi1 that Mel18 and Bmi1 act in synergy and in a dose-dependent and cell type-specific manner to repress Hox cluster genes and mediate cell survival of embryos during development. In addition, we demonstrate that Mel18 and Bmi1, although essential for maintenance of the appropriate expression domains of Hox cluster genes, are not required for the initial establishment of Hox gene expression. Furthermore, we show an unexpected requirement for Mel18 and Bmi1 gene products to maintain stable expression of Hox cluster genes in regions caudal to the prospective anterior expression boundaries during subsequent development.

Animals↗

Integration of Morphological and Genome-Wide SNP Data Reveals Regional Diversity in Thai Swamp Buffalo.

Thai swamp buffalo (Bubalus bubalis) are valuable animal genetic resources, but their regional diversity remains incompletely characterized. Morphological records were obtained from 799 buffaloes, and 474 individuals were genotyped using the 90K Axiom® Buffalo SNP Genotyping Array (Thermo Fisher Scientific, Applied Biosystems™, Santa Clara, CA, USA). Qualitative traits included coat color, horn shape, chevron pattern, and hair-whorl distribution, while quantitative characterization covered 30 body measurements. After excluding missing traits, 30 traits adjusted for sex, age, province, and farm from 768 animals were used for morphology-based analyses, which identified regional phenotypic differences but also substantial overlap, consistent with the influence of feeding, management, environment, and local selection on body conformation. Genome-wide SNP analyses indicated a broadly shared genetic background with detectable regional structure, and runs of homozygosity revealed regional differences in genomic autozygosity. The Lower South showed a comparatively strong recurrent-ROH pattern, whereas inference for the Upper South requires caution because of its small genomic sample. Overall, integrating morphology and genome-wide SNP data improves regional characterization, but data-dependent statistics, population-structure estimates, and conservation implications should be interpreted while regarding small sampling and unbalanced data.

Bubalus bubalis↗

Juvenile Pearson syndrome.

The clinical and magnetic resonance imaging findings of a 14-year-old boy with Pearson syndrome are presented. The patient represents the oldest living survivor of the original four patients described by Pearson and associates. This syndrome has recently been found to be associated with an mtDNA deletion. The patient reported here has a deletion similar but not identical to that reported in the literature. Several mitochondrial myopathies have been associated with mtDNA deletions, with considerable overlap between and among the phenotypes and underlying mtDNA deletions. The same may well prove to be true for Pearson syndrome.

Adolescent↗

Tumor variants by hormone receptor expression in white patients with node-negative breast cancer from the surveillance, epidemiology, and end results database.

PURPOSE: Hormone receptor expression (presence-positive or absence-negative) may reflect different stages of one disease or different breast cancer types. Determining whether hormone receptor expression represents one or more breast cancer phenotypes would have important paradigmatic and practical implications. METHODS: Breast cancer records were obtained from the National Cancer Institute's Surveillance, Epidemiology, and End Results (SEER) database. The study included 19,541 non-Hispanic white women with node-negative breast cancer. Standard tumor cell characteristics and breast cancer-specific survival were analyzed by independent estrogen receptor (ER+ and ER-), independent progesterone receptor (PR+ and PR-), and joint ERPR expression (ER+PR+, ER+PR-, ER-PR+, and ER-PR-). RESULTS: Age frequency density plots by hormone receptor expression showed two overlapping breast cancer populations with early-onset and/or late-onset etiologies. Independent ER+ and PR+ phenotype were associated with smaller tumor sizes, better grade, and better cancer-specific survival than ER- and PR- breast cancer types. Joint ERPR phenotype exhibited biologic gradients for tumor size, grade, and cancer-specific survival, which ranked from good to worse for ER+PR+ to ER+PR- to ER-PR+ to ER-PR-. CONCLUSION: Variations of standard tumor cell characteristics and breast cancer-specific survival by hormone receptor expression in white patients with node-negative breast cancer suggested two breast cancer phenotypes with overlapping etiologies and distinct clinical features.

Adult↗

Sexual selection on multivariate phenotype in wild and mass-reared Ceratitis capitata (Diptera: Tephritidae).

Males with a larger thorax and narrower face were found to be favoured by directional sexual selection in a previous field cage experiment where Ceratitis capitata males from the Seib 6-96 genetic sexing strain competed with wild males from Alto Valle (Patagonia) for the possession of wild females. Targets of sexual selection, however, might differ between wild and laboratory females as a response to adaptation to mass-rearing conditions. To evaluate possible divergences on the targets of sexual selection as a by-product of adaptation to mass-rearing conditions, field cage tests were performed with both wild and laboratory females. To avoid possible bias due to correlation among the measured traits (eye length [EL], face width [FW], head width [HW], and thorax length [TL]), a multivariate analysis was applied. Consistent with the previous experiment, the results indicated that TL and FW are probable targets of directional sexual selection independently of female strain. However, laboratory females were less selective than wild ones. Additionally, correlational sexual selection was detected acting on the multivariate phenotype. The effects of correlational selection overlap with those of directional selection on each single trait. The analysis of mating pair characteristics showed patterns that do not match the expectations for a random mating system. The current analysis indicates that during mating pair formation two processes overlap. On the one hand, sexual selection favours males with larger size (TL) and narrower faces (FW). This effect occurs in both wild and laboratory females. In addition, assortative mating based on both phenotype and origin was also observed.

Animals↗

Cloning and analysis of the mouse Fanconi anemia group A cDNA and an overlapping penta zinc finger cDNA.

Despite the cloning of four disease-associated genes for Fanconi anemia (FA), the molecular pathogenesis of FA remains largely unknown. To study FA complementation group A using the mouse as a model system, we cloned and characterized the mouse homolog of the human FANCA cDNA. The mouse cDNA (Fanca) encodes a 161-kDa protein that shares 65% amino acid sequence identity with human FANCA. Fanca is located at the distal region of mouse chromosome 8 and has a ubiquitous pattern of expression in embryonic and adult tissues. Expression of the mouse cDNA in human FA-A cells restores the cellular drug sensitivity to normal levels. Thus, the expression pattern, protein structure, chromosomal location, and function of FANCA are conserved in the mouse. We also isolated a novel zinc finger protein, Zfp276, which has five C(2)H(2) domains. Interestingly, Zfp276 is situated in the Fanca locus, and the 3'UTR of its cDNA overlaps with the last four exons of Fanca in a tail-to-tail manner. Zfp276 is expressed in the same tissues as Fanca, but does not complement the mitomycin C (MMC)-sensitive phenotype of FA-A cells. The overlapping genomic organization between Zfp276 and Fanca may have relevance to the disease phenotype of FA.

Adult↗

[Overlap of Sjögren-lupus erythematosus syndrome].

Patients with anti-SSA(Ro)-positive Sjögren's syndrome (SS)/lupus erythematosus (LE) overlap are a immunogenetically, serologically and clinically homogeneous group; what they have in common is an increased frequency of the HLA-DR3 phenotype, demonstrating SSA(Ro) antibody activity and the typical annular, polycyclic, erythematous lesions of subacute cutaneous LE. The sicca symptoms of SS may develop long after the cutaneous lesions of LE have been present, or vice versa, as well as vasculitic, purpuric, and Sweet's syndrome-like lesions. The elderly, predominantly female patients are at enhanced risk for pulmonary and neurological disease, but glomerulonephritis occurs infrequently. We present an 81-year-old, SSA(Ro) antibody-positive and HLA-DR3-positive woman with SS/LE overlap syndrome, who had cutaneous LE for 30 years, developing sicca symptoms, vasculitic and Sweet's syndrome-like skin lesions only years after the LE symptoms. Although there is still some discussion as to whether phenotypical SS/LE overlap represents a distinct disease or is secondary SS in SLE or systemic SS with cutaneous involvement, the SS/LE overlap syndrome seems well enough delineated to be considered as a disease entity with separate implications for therapy and prognosis. Its position in the spectrum of anti-SSA(Ro) antibody- and HLA-DR3-positive diseases and its relationship to SS and to anti-SSA(Ro) antibody-positive LE are discussed.

Aged↗

GATA3 haplo-insufficiency causes human HDR syndrome.

Terminal deletions of chromosome 10p result in a DiGeorge-like phenotype that includes hypoparathyroidism, heart defects, immune deficiency, deafness and renal malformations. Studies in patients with 10p deletions have defined two non-overlapping regions that contribute to this complex phenotype. These are the DiGeorge critical region II (refs 1, 2), which is located on 10p13-14, and the region for the hypoparathyroidism, sensorineural deafness, renal anomaly (HDR) syndrome (Mendelian Inheritance in Man number 146255), which is located more telomeric (10p14-10pter). We have performed deletion-mapping studies in two HDR patients, and here we define a critical 200-kilobase region which contains the GATA3 gene. This gene belongs to a family of zinc-finger transcription factors that are involved in vertebrate embryonic development. Investigation for GATA3 mutations in three other HDR probands identified one nonsense mutation and two intragenic deletions that predicted a loss of function, as confirmed by absence of DNA binding by the mutant GATA3 protein. These results show that GATA3 is essential in the embryonic development of the parathyroids, auditory system and kidneys, and indicate that other GATA family members may be involved in the aetiology of human malformations.

Amino Acid Sequence↗

Split hand/split foot deformity and LADD syndrome in a family: overlap between the EEC and LADD syndromes.

A mother and daughter are reported with apparently dissimilar syndromes. The mother has a split hand/split foot deformity and the daughter a condition consistent with a diagnosis of LADD syndrome. Absence of clefting and deficient formation of saliva and tears are the main signs that differentiate the LADD from the EEC syndrome. However, no distinct feature is constant between these two autosomal dominant disorders that show great phenotypic variability. This report emphasises the overlap between the LADD and the EEC syndromes.

Abnormalities, Multiple↗

Genetic diversity among barley cultivars assessed by sequence-specific amplification polymorphism.

We analyzed the genetic structure and relationships among barley cultivars (Hordeum vulgare L.) with sequence-specific amplification polymorphisms (S-SAPs). Polymorphisms were identified in 824 individual barley plants representing 103 cultivars (eight plants per cultivar) widely grown in Canada and the United States, using PCR primers designed from the long terminal repeat of the barley retrotransposon BARE-1 and a subset of four selective MseI primers. From the 404 bands scored, 150 were polymorphic either within or between cultivars. Genetic structure assessed with analysis of molecular variance attributed the largest component of variation to the within groups of cultivars (69-86%). Within-cultivar genetic variation was estimated as average gene diversity over loci and ranged from 0 (completely homogenous) to 0.076 (most heterogeneous cultivar). Only 17 out of 103 cultivars (16%) were judged to be homogenous by this criterion. Relationships among cultivars were analyzed by cluster analysis using unweighted pair-groups using arithmetic averages and found groups similar to those determined by agriculturally significant phenotypic traits such as spike morphology (two-rowed or six-rowed), cultivar type (malting or feed), seed characteristic (hull-less or hulled), and growth habit (winter or spring), with minor overlaps. Discriminant analysis of groups determined by these phenotypic traits fully supported the different groups with minor overlaps between the malting/feed. S-SAP markers generated from retrotransposons such as BARE-1 are invaluable tools for the study of genetic diversity in organisms with a narrow genetic base such as barley. In this study, S-SAP analysis revealed significant amounts of cryptic variation in closely related cultivars including somaclonal variation, which could not be inferred by the pedigree analysis.

Analysis of Variance↗