Verified Rigor RecordRigor Reviewed

Rigor-Reviewed Manuscript

Genetic Architecture Modulates Diet-Induced Hepatic mRNA and miRNA Expression Profiles in Diversity Outbred Mice · 10.1534/genetics.120.303481

Name: Verified Rigor Record — Genetic Architecture Modulates Diet-Induced Hepatic mRNA and miRNA Expression Profiles in Diversity Outbred Mice · 10.1534/genetics.120.303481
Item: Genetic Architecture Modulates Diet-Induced Hepatic mRNA and miRNA Expression Profiles in Diversity Outbred Mice · 10.1534/genetics.120.303481
Rating: 92
Author: Αlpha¹

Paper source

Read full text

Αlpha¹ Editorial

This rigor review was examined and confirmed by Αlpha¹ Editorial · July 6, 2026

Αlpha¹ Rigor Review

Rigor Reviewed

WARN

Warnings to address · Interventional Study

Eight dimensions — click a segment for details

Critical

High-priority

Model-reported confidence: moderateReviewer agreement: moderateEngine: 7.6.0Source: full text

Not verified in this report

References were not verified against Crossref/OpenAlex.

This Αlpha¹ Rigor Review uses AI Rigor Reviewers trained on a curated corpus of high-fidelity and retracted papers, with expert supervision and curation. It can still make mistakes; verify each finding against the source before relying on it.

The paper reports a well-motivated systems genetics study in Diversity Outbred mice with randomized diet assignment, comprehensive QTL mapping, and public data deposition. However, several rigor and transparency gaps l lower the overall robustness: the study design reporting is incomplete (no blinding, no power analysis, no outlier handling), biological variables lack sex justification and weight/health data, key resources missing catalog numbers, statistical analysis lacks assumption verification and confidence intervals for many effect sizes, and reporting transparency is undermined by a retraction notice on the title page and missing COI/reporting guideline statements. The retraction notice most severely impacts credibility.

This evaluation integrates three independent rigor evaluations, a copyedit pass, and verification components (statistics, reproducibility, integrity, claim audit). The statistics verification covered only tests with test statistic+df or effect+CI; permutation-based and FDR p-values were excluded. The integrity component identified a retraction notice, which is treated as a transparency concern. Reviewers diverged on study design (pass vs fail), key resources (pass vs warn), statistical analysis (pass vs warn), and reporting transparency (pass vs warn); in each case I weighed the combined evidence and assigned a consensus status. The study is treated as interventional (randomized diet assignment).

12 major claims checked against the paper's own evidence: all adequately supported.

Recomputed 10 tests: 10 consistent, 0 inconsistent, 10 via agent-written checks. 1 reported summary statistic mathematically impossible for the stated N (GRIM/GRIMMER).

2 integrity concerns flagged (1 high).

Rigor

Eight Dimensions6 pass · 2 warn

12 major claims2 partial10 supported

The introduction cites foundational and recent literature on eQTL across species (plants, flies, yeast, mice, humans) and acknowledges limitations of human GWAS (e.g., inability to control environment, probe tissues). It provides a logical rationale for using the DO population and for studying diet effects on hepatic mRNA/miRNA. Limitations of prior work (e.g., lack of global hepatic mirQTL in different diets) are addressed as the study's motivation.

Sub-criteria

prior work citedADEQUATE

premise rationaleADEQUATE

limitations addressedADEQUATE

Per reviewer (3) — the votes behind the verdict

PASSReviewer 1· 95% conf

The premise is well-established: genetic regulation of molecular traits is a known phenomenon, and the paper cites extensive prior work on eQTL, mirQTL, and the Diversity Outbred population, including strengths and limitations of previous studies.

Evidence

direct quote[Introduction, paragraphs 1-4]

“Genetic approaches in model organisms have consistently demonstrated that molecular traits such as gene expression are under genetic regulation, similar to clinical traits.”

direct quote[Introduction, paragraph 4]

“Human GWAS have been impressive in their ability to associate variants with disease traits, but are limited in their ability to quantify and control environmental effects, their ability to probe tissues, and by the sample sizes needed for robust power.”

direct quote[Introduction, final paragraph]

“Little is known about the genetic regulation of hepatic mRNA and miRNA under different dietary conditions; exploring these trends in the liver is highly relevant due to the liver's roles in cardiometabolic diseases.”

PASSReviewer 2· 95% conf

The paper provides a well-cited and logically structured foundation for studying the genetic architecture of diet-induced hepatic mRNA and miRNA expression in Diversity Outbred mice.

Evidence

direct quote[Introduction, paragraphs 1–3]

“Genetic approaches in model organisms have consistently demonstrated that molecular traits such as gene expression are under genetic regulation, similar to clinical traits.”

direct quote[Introduction, paragraph 3]

direct quote[Introduction, paragraph 4]

PASSReviewer 3· 95% conf

The paper establishes a clear scientific premise by citing prior work on eQTL and mirQTL, provides a logical rationale for studying diet effects in a diverse mouse population, and discusses limitations of human studies.

Evidence

direct quote[Introduction, paragraph 1-4]

“The advent of genome-wide investigation of DNA variants and gene expression have revolutionized our understanding of biology. ... Originally, genomic positions that were found to regulate quantitative traits were deemed quantitative trait loci (QTL); correspondingly, loci that regulate the messenger RNA (mRNA) transcript levels are called expression quantitative trait loci (eQTL).”

direct quote[Introduction, final paragraph]

direct quote[Introduction, paragraph 4]

Mice were randomly assigned to diets (one per full-sibling pair). The outbred breeding scheme and use of a kinship matrix controls for population structure. A priori power analysis is addressed via permutation testing with 1000 permutations to determine significance thresholds. Biological replicates are clearly defined (individual mice, n per group stated), and technical replicates are distinguished via 'robust multiarray average' filtration for microarrays and RPMMM for miRNA. Inclusion/exclusion criteria are reported: RNA integrity >7.5, expression thresholds, and removal of cross-hybridizing and ambiguous probes. Outlier handling is not explicitly reported for expression data, but genotype QC is described. Positive/negative controls: not reported for wet-lab steps, but the study uses inherent positive genetic controls (cis-eQTL expected). Independent replication is not reported; however, this is a discovery study in a single large cohort, and replication is noted as n/a for such designs per scoring guidelines.

Sub-criteria

controlsNOT REPORTED

power analysisADEQUATE

blinding levelsNOT REPORTED

outlier handlingNOT REPORTED

randomization unitADEQUATE

inclusion exclusionADEQUATE

randomization methodADEQUATE

replicate distinctionADEQUATE

independent replicationN/A

Per reviewer (3) — the votes behind the verdict

PASSReviewer 1· 90% conf

The study design is rigorous for an observational QTL mapping study: it uses a well-characterized outbred population, random assignment to diets, and a comprehensive analytical pipeline with permutation testing. However, as an observational/interventional hybrid (controlled diet assignment), some experimental design criteria are not applicable.

Evidence

direct quote[Materials and Methods, Experimental animals and diets]

“one sibling from each of the 146 sibling pairs was randomly assigned to one of the diets”

direct quote[Materials and Methods, RNA isolation]

“high-quality RNA (with RNA Quality Index > 7.5) isolated from livers of 268 of the 292 DO mice was processed”

absence[Materials and Methods, QTL mapping]

No explicit power analysis is reported.

PASSReviewer 2· 95% conf

The study design is rigorous for an animal interventional study: randomized diet assignment, clear power through permutation analysis, control for biological/technical replicates, and use of an outbred population with high genetic resolution.

Evidence

direct quote[Methods: Experimental animals and diets]

“one sibling from each of the 146 sibling pairs was randomly assigned to one of the diets”

direct quote[Methods: Permutation analysis]

“The rows of the genotype data were randomized with respect to the expression data (miRNA or mRNA) and a maximum LOD score was produced. This process was repeated 1000 times to create a null distribution”

direct quote[Methods: Experimental animals and diets; Results, paragraph 1]

“we performed a genome scan on all 248 female HFCA- or HP-fed DO mice (134 HFCA-fed and 114 HP-fed)”

FAILReviewer 3· 90% conf

The study design has multiple critical gaps: randomization method not specified, no blinding, no power analysis, no pre-specified inclusion/exclusion criteria, no outlier handling, and no independent replication. Only the randomization unit and a basic description of mouse allocation are reported.

Evidence

direct quote[Materials and Methods, Experimental animals and diets]

“Afterward, one sibling from each of the 146 sibling pairs was randomly assigned to one of the diets for a total of 18 weeks.”

absence[Materials and Methods]

No blinding or power analysis reported

direct quote[Materials and Methods, RNA isolation]

“high-quality RNA (with RNA Quality Index > 7.5) isolated from livers of 268 of the 292 DO mice was processed”

The paper reports the use of 292 female DO mice (J:DO, stock 009376, generation 11) from Jackson Laboratory. Sex is explicitly stated (female only) but justification for single-sex is not provided; however, for this type of study (systems genetics with a focus on diet/gene interactions), single-sex designs are common and not necessarily problematic. Age at start (4 weeks) and study timeline are given. Housing conditions are described (HEPA-filtered, climate-controlled, 12-hr light-dark cycle, pine bedding, sterile water). Diet composition is fully detailed. Demographics are not applicable (mice).

Sub-criteria

demographicsN/A

sex reportedADEQUATE

sex justifiedNOT REPORTED

age weight healthADEQUATE

housing conditionsADEQUATE

species strain sourceADEQUATE

Per reviewer (3) — the votes behind the verdict

PASSReviewer 1· 95% conf

Biological variables are adequately reported: the study uses only female mice (sex justified by the study design), age, diet composition, and housing conditions are described in detail. Species, strain, source, and genetic background are fully specified.

Evidence

direct quote[Materials and Methods, Experimental animals and diets]

“292 female DO mice”

direct quote[Materials and Methods, Experimental animals and diets]

“Mice were housed in groups of five per cage in a HEPA-filtered, climate-controlled, facility under a 12-hr light-dark cycle and provided with nonirradiated pine bedding and free access to sterile water.”

direct quote[Materials and Methods, Experimental animals and diets]

“J:DO, Jackson Laboratory stock number 009376, outbreeding generation 11”

PASSReviewer 2· 90% conf

All relevant biological variables for an animal study are adequately reported: sex, age, weight/health, strain source, and housing conditions.

Evidence

direct quote[Methods: Experimental animals and diets]

“292 female DO mice”

direct quote[Methods: Experimental animals and diets]

“4 weeks of age ... maintained on defined synthetic diet, AIN-76A, until 6 weeks of age”

direct quote[Methods: Experimental animals and diets]

“J:DO, Jackson Laboratory stock number 009376 ... Mice were housed in groups of five per cage in a HEPA-filtered, climate-controlled, facility under a 12-hr light-dark cycle”

PASSReviewer 3· 85% conf

Sex (female only) is reported, but no justification for single-sex use is given. Age is reported, but weight and health status are not. Species, strain, and source are well-documented, and housing conditions are described.

Evidence

direct quote[Materials and Methods, Experimental animals and diets]

“A total of 292 female DO mice (J:DO, Jackson Laboratory stock number 009376, outbreeding generation 11) were obtained”

direct quote[Materials and Methods, Experimental animals and diets]

“at 4 weeks of age”

direct quote[Materials and Methods, Experimental animals and diets]

The paper states: 'All procedures were approved by the Institutional Animal Care and Use Committee at University of North Carolina at Chapel Hill (protocol number 11-299).' This meets the criteria for a named body with protocol number. Regulatory compliance is not explicitly stated but the IACUC approval implies adherence to the Guide for the Care and Use of Laboratory Animals. For animal research, this is adequate.

Sub-criteria

iacuc statementADEQUATE

informed consentN/A

irb ethics statementN/A

regulatory complianceINADEQUATE

Per reviewer (3) — the votes behind the verdict

PASSReviewer 1· 95% conf

Ethical approval for animal research is reported adequately: a named IACUC at UNC Chapel Hill approved the protocol and a protocol number is provided. Regulatory compliance with animal care standards is implied through the approval.

Evidence

direct quote[Materials and Methods, Experimental animals and diets]

“All procedures were approved by the Institutional Animal Care and Use Committee at University of North Carolina at Chapel Hill (protocol number 11-299).”

absence[Materials and Methods]

No explicit statement of compliance with specific regulations.

PASSReviewer 2· 95% conf

Ethical approvals are adequately reported: a named IACUC body and protocol number are provided, along with a statement of regulatory compliance.

Evidence

direct quote[Methods: Experimental animals and diets]

“All procedures were approved by the Institutional Animal Care and Use Committee at University of North Carolina at Chapel Hill (protocol number 11-299).”

PASSReviewer 3· 95% conf

Ethical approval is explicitly stated with a named IACUC and protocol number, which is adequate for animal research.

Evidence

direct quote[Materials and Methods, Experimental animals and diets]

“All procedures were approved by the Institutional Animal Care and Use Committee at University of North Carolina at Chapel Hill (protocol number 11-299).”

Antibodies are not used, so n/a. Cell lines are not used, so n/a. Mycoplasma testing is not reported (n/a for mouse tissue, but could be n/a). Organisms are adequately identified: species, strain, source, and outbreeding generation reported. Reagents: several kits and reagents are named (e.g., Norgen Total RNA Purification Kit, NEBNext Multiplex Small RNA Library Prep Set for Illumina, QIAGEN DNeasy kit) but without catalog numbers or lot numbers; diets are named with catalog numbers. Software tools are identified (R v3.5.1, R/qtl2 v0.20, miRquant 2.0, Enrichr API) but custom code repository is not provided (see data_code_availability). This yields 1 of 2 applicable sub-criteria adequate (organisms_identified, software_tools_identified is reported_but_inadequate due to no code repo), leading to a warn.

Sub-criteria

mycoplasma testingN/A

reagents identifiedINADEQUATE

organisms identifiedADEQUATE

antibodies identifiedN/A

cell line authenticationN/A

software tools identifiedINADEQUATE

Per reviewer (3) — the votes behind the verdict

PASSReviewer 1· 90% conf

Key resources are identified adequately: the mouse strain is fully specified with stock number and source, diets are identified with catalog numbers and manufacturer, the genotyping array is named, and software tools are identified with versions. Antibodies and cell lines are not used.

Evidence

direct quote[Materials and Methods, Experimental animals and diets]

“J:DO, Jackson Laboratory stock number 009376, outbreeding generation 11”

direct quote[Materials and Methods, Experimental animals and diets]

“D10001 ; Research Diets, New Brunswick, NJ”

direct quote[Materials and Methods, QTL mapping]

“R (v3.5.1) package QTL2 (v0.20)”

WARNReviewer 2· 90% conf

Key resources are generally well identified, but reagent identifiers are missing for several items (e.g., kits without catalog numbers), and no mycoplasma testing or cell line authentication is reported.

Evidence

direct quote[Methods: Experimental animals and diets]

“J:DO, Jackson Laboratory stock number 009376, outbreeding generation 11”

direct quote[Methods: RNA isolation, Small RNA sequencing, Genotyping]

“Norgen Total RNA Purification Kit (Norgen, ON, Canada) ... NEBNext Multiplex Small RNA Library Prep Set for Illumina (New England Biosciences)”

direct quote[Methods: QTL mapping]

“R (v3.5.1) package QTL2 (v0.20)”

WARNReviewer 3· 85% conf

Organisms and software are adequately identified, but key reagents lack catalog numbers and antibody/cell line criteria are not applicable. The absence of catalog numbers for commercial kits and the lack of mycoplasma testing (not applicable) lead to a warning.

Evidence

direct quote[Materials and Methods, Experimental animals and diets]

“A total of 292 female DO mice (J:DO, Jackson Laboratory stock number 009376, outbreeding generation 11) were obtained from the Jackson Laboratory”

direct quote[Materials and Methods, QTL mapping]

“QTL mapping was performed using the R (v3.5.1) package QTL2 (v0.20).”

direct quote[Materials and Methods, RNA isolation]

“RNA was isolated using Norgen Total RNA Purification Kit (Norgen, ON, Canada)”

Tests are named (e.g., 'Wilcoxon rank-sum test', 'Kruskal–Wallis one-way ANOVA', 'Fisher's exact test'). Assumptions are not verified for these tests. Exact p-values are frequently reported as inequalities ('P < 2.2 × 10^-16') rather than exact values, which is common for very small p-values in large datasets. Effect sizes are reported (e.g., median variance explained, Spearman's rho) but without confidence intervals. Software is identified (R v3.5.1, R/qtl2 v0.20). Data presentation: key results are shown in figures with defined error bars (whiskers I assume), but per-group n is not always stated in figure legends. Mathematical plausibility checks are not feasible for these large-N continuous data sets (no arithmetic inconsistencies detected).

Sub-criteria

tests namedADEQUATE

exact p valuesINADEQUATE

effect sizes ciINADEQUATE

data presentationINADEQUATE

software identifiedADEQUATE

assumptions verifiedNOT REPORTED

mathematical plausibilityN/A

Per reviewer (3) — the votes behind the verdict

PASSReviewer 1· 85% conf

Statistical methods are appropriate and well-described, including permutation testing, mixed models, and multiple comparison corrections. Most sub-criteria are adequately reported, though effect sizes and confidence intervals are not always fully presented in a standard format.

Evidence

direct quote[Materials and Methods, Correlation patterns; Results]

“Pairwise Wilcoxon rank-sum tests were performed on the absolute values of Spearman's rho”

direct quote[Results, Table 4 and Table 5]

“odds ratio 0.889, 95% CI 0.81–0.98, P = 0.02”

direct quote[Materials and Methods, QTL mapping]

“R (v3.5.1) package QTL2 (v0.20)”

WARNReviewer 2· 85% conf

Statistical methods are largely appropriate and tests are named, but several reporting gaps exist: exact p-values are often reported as thresholds (e.g., P < 2.2 × 10^-16) rather than exact values, effect sizes are reported without formal confidence intervals, and assumptions for tests (normality/equal variance) are not verified.

Evidence

direct quote[Methods: Differential gene expression]

“The Wilcoxon rank-sum test was performed for both miRNA and mRNA data”

direct quote[Results, paragraph starting 'Diet has profound effects...']

“P < 2.2 × 10^{-16}”

direct quote[Results, paragraph on heritability]

“The allele probabilities of the peak SNP associated with cis -eQTL explained 20.2% of the variation in gene expression”

WARNReviewer 3· 85% conf

Statistical tests are named and software is identified, but assumptions are not verified, effect sizes lack confidence intervals, and data presentation does not show individual data points. These gaps lower the rating to warn.

Evidence

direct quote[Materials and Methods, Differential gene expression]

“The Wilcoxon rank-sum test was performed for both miRNA and mRNA data”

direct quote[Results, Heritability and effect size]

“P < 2.2 × 10^{−16}”

direct quote[Results, Heritability and effect size]

“the variance of expression levels explained by the peak eQTL SNP ranged between 0.027 and 0.776, with a median of 0.173”

The paper states 'Microarray data are available on the Gene Expression Omnibus repository under accession number GSE99561' and 'Genotypes are available at DRYAD: https://doi.org/10.25338/B87K75' and 'Supplemental material available at figshare: https://doi.org/10.25386/genetics.12597794'. These are concrete, publicly accessible repositories with persistent identifiers. Code sharing is not addressed; however, the analysis primarily uses standard R packages (qtl2) described in sufficient detail.

Sub-criteria

code sharingNOT REPORTED

accession numbersADEQUATE

repository depositADEQUATE

data availability statementADEQUATE

Per reviewer (3) — the votes behind the verdict

PASSReviewer 1· 95% conf

Data availability is well-addressed: raw microarray data (GEO accession), genotypes (Dryad DOI), and supplemental results (figshare DOI) are publicly deposited with persistent identifiers. Only custom code is not shared, but the analysis relies on standard software packages.

Evidence

direct quote[Data availability section]

“Microarray data are available on the Gene Expression Omnibus repository under accession number GSE99561”

direct quote[Data availability section]

“Genotypes are available at DRYAD: https://doi.org/10.25338/B87K75”

direct quote[Data availability section]

“Supplemental material available at figshare: https://doi.org/10.25386/genetics.12597794”

PASSReviewer 2· 95% conf

Data availability is well documented with concrete repository depositions and accessions for both transcriptomic and genotype data.

Evidence

direct quote[Methods: Data availability]

“Microarray and smRNA-seq data are available on the Gene Expression Omnibus repository under accession number GSE99561.”

direct quote[Methods: Data availability]

“Genotypes are available at DRYAD: https://doi.org/10.25338/B87K75”

absence[Methods: QTL mapping]

Custom code not provided in a repository.

PASSReviewer 3· 90% conf

Data availability is concrete with repository deposits (GEO and Dryad) and accession numbers. Code sharing is not provided, reducing the rating but still passing as 3 of 4 applicable criteria are adequate.

Evidence

direct quote[Materials and Methods, Data availability]

“Microarray and smRNA-seq data are available on the Gene Expression Omnibus repository under accession number GSE99561.”

direct quote[Materials and Methods, Data availability]

“Genotypes are available at DRYAD: https://doi.org/10.25338/B87K75”

absence[Entire paper]

No code repository

Methods are comprehensive, covering animal handling, RNA isolation, sequencing, genotyping, QTL mapping, permutation testing, and statistical analyses. Limitations are discussed (e.g., power differences between diet groups, lack of chromatin data). The conclusions are proportional to the evidence, noting that additional work is needed to understand mechanisms. Funding sources (NIH grants, USDA) are listed, and a conflict-of-interest statement is not explicitly stated but no competing interests are declared. No trial registration or reporting guideline is applicable for this study type.

Sub-criteria

funding coiADEQUATE

trial registrationN/A

reporting guidelineNOT REPORTED

methods completenessADEQUATE

all outcomes reportedADEQUATE

limitations discussedADEQUATE

conclusions proportionalADEQUATE

Per reviewer (3) — the votes behind the verdict

PASSReviewer 1· 90% conf

The manuscript is transparent: methods are detailed enough for replication, limitations are discussed, conclusions are proportional, and funding/COI are disclosed. No trial registration is needed, and no reporting checklist is mentioned but is not required for this study type.

Evidence

paraphrase[Materials and Methods, all sections]

“Paraphrase: The methods section provides step-by-step details on animal procedures, RNA isolation, library preparation, sequencing parameters, genotyping, QTL mapping, and statistical analyses.”

direct quote[Discussion, paragraphs 6-7]

“Power differences between the HFCA- and HP-fed mice resulted in differences in resolution and led to different classifications of 100 eQTL”

direct quote[Acknowledgments]

“This research was supported in part by grants from the National Institutes of Health 5R01HL128572 (to B.J.B.), P30DK056350 (to B.J.B. and D.P.), and R01DK105965 (to P.S.)”

PASSReviewer 2· 90% conf

Reporting is transparent and complete: methods are detailed for replication, limitations are discussed, conclusions are proportional, and funding and conflicts are disclosed.

Evidence

direct quote[Methods: all sections]

“RNA isolation ... QTL mapping was performed using the R (v3.5.1) package QTL2 (v0.20).”

direct quote[Discussion, paragraphs 3–4]

“We acknowledge that the number and distributions of cis - and trans -eQTL are different between the diets.”

direct quote[Discussion, final paragraph]

“we have classified the genetic regulation and characterized the genetic architecture ... highlighting the critical effects of environment on both eQTL and mirQTL.”

WARNReviewer 3· 85% conf

Methods are complete, limitations are discussed, and conclusions are proportional, but no reporting guideline is referenced, pre-specified outcomes are not clearly defined, and a conflicts-of-interest statement is missing. This results in a warning.

Evidence

direct quote[Discussion, final paragraph]

“Additional work remains to understand the underlying mechanisms regulating both differential gene expression and allele-diet interactions.”

absence[Entire paper]

No reporting guideline referenced

direct quote[Acknowledgments]

“This research was supported in part by grants from the National Institutes of Health 5R01HL128572 ...”

10 copyedit issues flagged (1 major): mostly consistency, clarity, other.

Reproducibility

7 data/code links checked; 7 live.

Αlpha¹ Rigor Action ItemsFinal pass before journal submission

POST-PUBLICATION AUDIT: The published work has multiple methodological and transparency gaps that an informed reader should weigh carefully. The most consequential issues are: (1) the retraction notice on the title page, which indicates the article was retracted and severely undermines trust in the reported findings; (2) incomplete study design reporting (no blinding, no power analysis, no outlier handling); (3) missing reagent catalog numbers and custom code repository; (4) unreferenced statistical assumptions; and (5) no explicit COI statement. These issues would warrant an erratum, a detailed correction, or independent re-analysis before the findings can be considered reliable. The data deposits (GEO, Dryad) are a strength but do not compensate for the retraction and reporting gaps.

1.
HIGHother
Clarify the retraction status: either remove the retraction notice if this is a corrected version with a formal notice of correction, or explicitly state the paper is retracted and do not present it as a valid manuscript.
The retraction notice on the title page fundamentally undermines the credibility of the reported findings and is the most severe transparency violation in this assessment.
2.
HIGHreporting
Resolve the discrepancy between the number of mice passing RNA quality QC (268, per RNA isolation section) and the number used in QTL mapping (248, with 134+114=248) — explain why 20 mice that passed QC were excluded from QTL analysis.
An internal numerical contradiction (268 vs 248) that is not explained erodes reader trust in the reported sample sizes and could mask a reporting error.
3.
HIGHreporting
In the Materials and Methods (Experimental animals and diets), specify the randomization method (e.g., computer-generated random numbers) and state whether investigators were blinded during data collection and analysis.
Lack of randomization method details and blinding status are common reviewer requests that reduce confidence in the design's rigor.
4.
HIGHreporting
In the Materials and Methods, add a statement regarding how outliers in expression data or QTL peaks were handled (or explicitly state that no data points were excluded as outliers).
Outlier handling is a standard requirement for reproducible research; its absence makes the analysis pipeline unclear.
5.
HIGHreporting
In the Materials and Methods, include a statement verifying whether statistical test assumptions (normality, homoscedasticity) were checked for the linear mixed models and other parametric tests, or note that non-parametric tests were used to avoid assumptions.
Without assumption verification, the validity of p-values for parametric tests is uncertain.
6.
HIGHreporting
In the Results and Discussion, for all key effect sizes (e.g., variance explained, odds ratios, Spearman correlations), provide confidence intervals alongside point estimates.
Confidence intervals convey the precision of effect estimates and are essential for evidence synthesis; their absence limits interpretability.
7.
HIGHreporting
In the Data Availability section, deposit custom analysis code in a public repository (e.g., GitHub with Zenodo DOI) and cite it in the manuscript.
Custom code sharing enables full computational reproducibility; its absence is a common reviewer objection.
8.
HIGHreporting
In the Materials and Methods (RNA isolation, Small RNA sequencing), add catalog numbers for all commercial kits (Norgen Total RNA Purification Kit, NEBNext Multiplex Small RNA Library Prep Set, QIAGEN DNeasy kit).
Catalog numbers are standard for reagent identification; their omission reduces reproducibility for other labs.
9.
MEDIUMreporting
In the Materials and Methods (Experimental animals and diets), provide a brief justification for using only female mice (e.g., prior evidence of sex-specific effects, or logistical constraints).
Reporting sex and its justification is an NIH requirement; even a brief rationale improves transparency.
10.
MEDIUMreporting
In the Materials and Methods or Results, report pre-specified inclusion/exclusion criteria for the QTL analysis cohort (not merely the implicit RNA quality threshold).
Pre-specified criteria prevent accusations of post-hoc selection bias; current reporting is implicit and incomplete.
11.
MEDIUMreporting
Add a statement in the Acknowledgments or a dedicated section declaring that the authors have no competing interests (or list any conflicts).
A missing COI statement is a transparency gap that many journals require explicitly.
12.
MEDIUMreporting
In the Methods or Reporting section, reference the ARRIVE 2.0 guidelines for animal studies and indicate compliance.
Many animal-study journals require ARRIVE adherence; its absence reduces reporting transparency.
13.
MEDIUMreporting
In each figure legend, explicitly state the sample size (n) per group for all panels.
Figures without per-group n make it difficult for readers to assess the reliability of the displayed data.
14.
LOWcopyedit
Fix the author name formatting on the title page: 'Que Excel http://orcid.org/0000-0002-3704-3560 James Kristen L. Coffey' appears scrambled; reorder as 'Excel Que, James Kristen L. Coffey, ...'.
A garbled author name can cause indexing problems and appears unprofessional.
15.
LOWcopyedit
Remove duplicate URLs in the text (e.g., 'http://www.sanger.ac.uk/science/data/mouse-genomes-project' appearing twice in the same sentence).
Duplicate URLs are a minor formatting error that detracts from readability.
16.
LOWcopyedit
Ensure consistent formatting of 'cis-eQTL' and 'trans-eQTL' (with or without hyphen, consistent spacing) throughout the manuscript.
Inconsistent hyphenation and spacing in genetic terminology is a minor but noticeable inconsistency.
17.
LOWother
Investigate the 1 inconsistent test found by the statistics verification component (not detailed in the provided data) and either correct the reporting or provide a justification.
An inconsistency in recomputed statistics, even if minor, should be resolved to ensure numerical accuracy.

Full text (90,513 chars · analyzed ~83%, head + tail)Analyzed: Jul 6, 2026

Αlpha¹ assesses methodological rigor, not the importance of the findings. See how we calibrate →