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Table 2 Comparison of methodological approaches for analysis of gene expression data

From: Use of gene expression studies to investigate the human immunological response to malaria infection

Dataset

Data generation

Gene ontology analysis

GEO series

Publication

RNA Quantification Platform

Normalization

Adjustment for covariates

Definition expression

Expressed genes

Threshold FC

Threshold P

Test

Multiple testing

GO analysis

Threshold GO enrichment p

Test

Multiple testing

GSE2900

Griffiths (2005)

Stanford University cDNA lymphochip two color microarray

Scaled to geometric mean of sample:reference signal ratio from all array features

NS

Signal threshold

9869

2.5 (from median in > 4 samples)

0.1

Permutation

FDR

NA

NA

NA

NA

GSE5418

Ockenhouse (2006)

Affymetrix U133A GeneChips

RMA

NS

NS

NS

No

0.01

SAM, t-test

FDR

Onto Express and Pathway Architect

0.05

NS

FDR

GSE15221

Franklin (2009) and Sharma (2011)

Illumina Human WG-6 v2.0

Cubic spline

NS

Signal threshold

NS

1.7

0.01

Paired t-test

FDR

Onto Express

Varying

NS

NS

GSE15221

Hirako (2018)

Illumina Human WG-6 v2.0

Cubic spline

NS

Signal threshold

NS

1.5

0.01

Permutation and t-test

FDR

DAVID, GSEA

0.05

Multiple

FDR

GSE26876

de Carvalho (2011)

Affymetrix Human Gene 1.0 ST Array

RMA

NS

NS

NS

1.5

0.05

Student t-test

No

Ingenuity pathway analysis

NS

NS

NS

GSE33811

Krupka (2012)

Affymetrix Human Gene 1.0 ST Array

RMA and Quantile

NS

Signal and variation threshold

3110

2

0.05

Paired t-test

No

Gene set enrichment analysis on selected GO terms

0.01

Paired t-test

FDR

GSE34404

Idaghdour (2012)

Illumina Human HT-12 BeadChips

Quantile

Location, Sex, Hb, total cell counts (RBCs and WBCs) and ancestry

Signal and normality threshold

NS

2 (for comparison)

0.01

ANOVA, ANCOVA

FDR

Gene set enrichment analysis on customized MsigDB database

0.05

NS

Bonferroni

GSE55843

Jagannathan (2014)

Agilent Sure Print G3 Human Gene Expression 8 × 60K v2 gene expression microarrays

Quantile

NS

Signal threshold

NS

2

0.05

SAM

FDR

NA

NA

NA

NA

GSE53292

Jaijyan (2015)

Illumina Genome Analyzer Iix 72SE

NS

NS

NS

NS

NS

0.05

t-test

No

GeneCodis3, Bingo 2.3 plugin (Cytoscape 2.8.3)

0.05

NS

NS

GSE50957 GSE52166

Tran (2016)

Illumina HiSeq 2000 2 × 100 PE

TAMM

Batch, Sex, Age, Pre-infection baseline

Signal and variation threshold, removal Y chromosomes

NS

1.5

0.05

Limma

FDR

Ingenuity pathway analysis

0.05

Fisher exact test

FDR

GSE50957 GSE67184

Vallejo (2018)

Illumina HiSeq 2000 2 × 100 PE

CPM, TPM

NS

Signal threshold

NS

NS

0.05

EdgeR

FDR

WGSEA, ToppGene, STRING

0.05

Multiple

FDR

GSE64338

Subramaniam (2015)

Affymetrix Human Gene 1.0 ST Array

Nonlinear normalization based on Li-Wong methods

NS

NS

NS

1.2

0.001

Paired t-test

FDR

Ingenuity Pathway Analysis

0.05

NS

FDR

GSE64493

Sullivan (2015)

Agilent Sure Print G3 Human Gene Expression 8 × 60K v2 gene expression microarrays

Quantile

NS

Signal threshold

NS

1.5

0.03

Limma

FDR

DAVID

0.05

NS

FDR

GSE67184

Rojas-Penas (2015)

Illumina HiSeq 2500 2 × 100 PE

SNM

Location/time-point, subject (random effect)

Signal threshold

6154

No

0.05

NS

FDR

NA

NA

NA

NA

GSE67184

Gardinassi (2018)

Illumina HiSeq 2500 2 × 100 PE

NS

NS

NS

NS

No

0.05

Limma, repeated measures ANOVA

FDR

GSEA on blood transcriptome modules (BTM, Li et al.)

0.05

permutation

FDR

GSE7586

Muehlenbachs (2007)

Affymetrix U133 Plus 2.0 GeneChip

GC RMA

NS

NS

NS

2.5

0.01

t-test

No

NA

NA

NA

NA

GSE77122

Tarawa (2017)

Agilent Sure Print G3 Human Gene Expression 8 × 60K gene expression microarrays

Each gene expression array dataset was normalized to the in silicon pool for the macrophages cultured with RBCs

NS

NS

NS

No

0.05

Paired t-test

No

DAVID

0.05

Fisher exact test

No

GSE93664

Burl (2017)

Affymetrix Human Gene ST 2.0 gene array

RMA

NS

NS

NS

2

0.05

NS

No

STRING

0.01

NS

Corrected unspecified

GSE100562

Quin (2017)

Illumina HiSeq 2500 2 × 50 PE

NS

NS

NS

NS

No

0.05

Limma

FDR

NA

NA

NA

NA

GSE1124

Boldt (2019)

Affymetrix U133A + B GeneChips

RMA

NS

Signal threshold

NS

1.9

0.004

SAM

FDR

DAVID and Ingenuity Pathway Analysis

0.05

NS

NS

GSE114076

Terkawi (2018)

Agilent Sure Print G3 Human Gene Expression 8 × 60K gene expression microarrays

Each gene expression array dataset was normalized to the in silicon pool for the neutrophils cultured with RBCs

NS

NS

NS

2

0.01

Limma

No

Genomatix GeneRanker, DAVID, NET-GE and Enricher

0.05

NS

Corrected unspecified

GSE97158

Rothan (2018)

Illumina HiSeq 2500 2 × 51 PE

TMM

Blocking by subject, in two separate models interaction with cell count and time of parasitemia was added

Signal threshold

16,473

1.5

0.05

Limma

FDR

GSEA (camera) on blood transcriptome modules (BTM, Li et al.)

0.05

Fisher exact test

FDR

GSE65928

Portugal (2015)

Affymetrix Human Gene ST 2.0 gene array

RMA

NS

NS

NS

NS

0.05

ANOVA

FDR

Ingenuity pathway analysis

NS

NS

NS

GSE72058

Feintuch (2016)

Affymetrix Human Gene 1.0 ST array

RMA and Quantile

Peripheral parasitemia

NS

NS

No

0.05

t-test

No

GSEA, CateGOrizer and ingenuity pathway analysis

0.2 and 0.06

NS

FDR

  1. FDR false discovery rate, Hb haemoglobin, NA not available, NS not specified in publication, RBCs red blood cells, RMA Robust Multichip average, SNM supervised normalization of microarray, TMM trimmed mean of M-values, GEO Gene Expression Omnibus, GE gene ontology