Table 1

Checklist for authors' of MIQE précis at time of manuscript submission, detailing information about individual parameters associated with each step of the RT-qPCR workflow.

Sample/Template

details


Source

If cancer, was biopsy screened for adjacent normal tissue?

Method of preservation

Liquid N2/RNAlater/formalin

Storage time (if appropriate)

If using samples >6 months old

Handling

fresh/frozen/formalin

Extraction method

TriZol/columns

RNA: DNA-free

Intron-spanning primers/no RT control

Concentration

Nanodrop/ribogreen/microfluidics

RNA: integrity

Microfluidics/3':5' assay

Inhibition-free

Method of testing


Assay optimisation/validation


Accession number

RefSeq XX_1234567

Amplicon details

exon location, amplicon size

Primer sequence

even if previously published

Probe sequence*

identify LNA or other substitutions

In silico

BLAST/Primer-BLAST/m-fold

empirical

primer concentration/annealing temperature

Priming conditions

oligo-dT/random/combination/target-specific

PCR efficiency

dilution curve

Linear dynamic range

spanning unknown targets

Limits of detection

LOD detection/accurate quantification

Intra-assay variation

copy numbers not Cq


RT/PCR


Protocols

detailed description, concentrations, volumes

Reagents

supplier, Lot number

Duplicate RT

ΔCq

NTC

Cq & melt curves

NAC

ΔCq beginning:end of qPCR

Positive control

inter-run calibrators


Data analysis


Specialist software

e.g., QBAsePlus

Statistical justification

e.g., biological replicates

Transparent, validated normalisation

e.g., GeNorm summary


*Disclosure of probe sequences is strongly encouraged.

Bustin et al. BMC Molecular Biology 2010 11:74   doi:10.1186/1471-2199-11-74

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