Checklist for the description of sequence variants

Since references to WWW-sites are not yet acknowledged as citations, please mention den Dunnen JT and Antonarakis SE (2000). Hum.Mutat. 15:7-12 when referring to these pages.

Purpose

Going through publications (JT den Dunnen, in prep.) one can easily see where people tend to offend the "Current recommendations for the description of sequence variants". The checklist below covers the most problematic issues and should assist those preparing a publication to describe sequence variants following the current recommendations.

Checklist

1. Reference Sequence - do you clearly describe the sequence used as a reference sequence ?
   A publication should mention, preferably in the Materials & Methods section and or Table legend, which sequence file was used as reference sequence for numbering of the residues (DNA, RNA and protein); see Recommendations, Discussion and mtDNA variants.
   • do you mention a GenBank (not GeneBank) RefSeq-file accession number with version number ?; do not forget the underscore in the accession number (correct is NM_004006.2, not NM004006.2).
   • for a coding DNA reference sequence, do you clearly state that nucleotide numbering uses the A of the ATG translation initiation start site as nucleotide +1 ?
   • a genomic reference sequence starts with nucleotide 1; a genomic reference sequence can thus not have negative numbers
2. Intronic variants - do you indicate where the reference intron sequence can be found ?
   the recommendation is to describe intronic variants in the format "c.89-2A>G" and not like "c.IVS4-2A>G" (see Discussion). When the format "c.IVS4-2A>G" is used, do you clearly indicate on which GenBank file intron / exon numbering is based and where the reference intron sequence can be found ?
3. Tabular overview - do you provide a clear, unequivocal overview of all changes reported ?
   preferably, a publication contains a tabular overview of all sequence changes reported. This overview contains columns describing the change at DNA-level (absolutely essential) and, optional, at RNA and protein level. When data on RNA and/or protein level are provided, it should be made clear whether the data were deduced or experimentally verified (e.g. state explicitly when RNA was analysed to confirm the putative splice mutation detected).
4. Insertions
   • are insertions reported in the format c.51_52insT ?
     since it is not clear whether one means insertion at or insertion after position 52, insertions should not be reported as c.52insT but in the format c.51_52insT (see Discussion).
   • are the insertions reported really insertions or are they in fact duplications ?
     duplicating insertions should be described as duplications, not as insertions; c.92_94dup (or c.92_94dupGAC) is correct, c.94_95insGAC is not correct (see Discussion). 
5. Most 3' position - do you correctly assign the change to the most 3' position possible ?
   for deletions, duplications and insertions the most 3' position possible is arbitrarily assigned to have been changed (see Recommendations); important especially in single residue (nucleotide or amino acid) stretches or tandem repeats. Example ACTTTGTGCC to ACTTGCC is described as c.5_7delTGT (not as c.4_6delTTG)
6. Recessive diseases - do you clearly describe which changes are found in which combination ?
   a publication describing sequence changes found in patients suffering from a recessive disease should for each patient explicitly mention which combination of (pathogenic) changes was identified (see Recommendations). Example c.[76C>T]+[87G>A] or c.[76C>T]+[?].
   NOTE: this description differs from that describing several changes in one allele, which has the format c.[76A>C; 113G>C].
7. Range - is the sign used to indicate a range a "_" (underscore) and not a "-" (minus) ?
   to prevent confusion, the underscore should be used to indicate a range and not the minus sign. The minus sign should only be used to indicate negative numbers. The correct description to indicate a deletion of the coding DNA nucleotides 12 to 14 is c.12_14del. Not correct is c.12-14del, which describes a deletion of nucleotide -14 in the intron directly preceding cDNA nucleotide 12 (see Discussion).
8. Deletion - do you indicate the first and last residue involved in a deletion ?
   a deletion of more than one residue should mention the first and last residue deleted, separated using a "_" (underscore). Example c.21_24del or p.Ala13_Gln16del.
9. Describe at DNA-level - do you describe all changes reported at DNA-level ?
   all changes reported must be described at DNA-level
   • when descriptions at RNA or protein level are given in the text, upon first appearance, use a format like "c.78G>C (p.Trp26Cys)"
   • description of "silent mutations" in the format "p.Leu54Leu (or p.L54L)" is not allowed. Descriptions should be given at DNA level, it is non-informative and not unequivocal (there are five possibilities at DNA level); a correct description is c.162C>G 
10. RNA protein level descriptions
    recommendations exist to describe alternative transcripts deriving from one allele (see Recommendations). Since these descriptions are rather complex to explain, it is wise to include a link to the HGVS recommendations in the publication.
11. Protein level descriptions
    • protein reference sequence - the protein reference sequence should represent the primary translation product, not a processed mature protein, and thus include any signal peptide sequences (see Recommendations).
    • one/three letter amino acid code - are the correct amino acid codes used at protein level ?;  several amino acids start with the same initial letter (Ala, Arg, Asn, Asp start with A, Gln, Glu, GLy with G, Leu, Lys with L, Phe, Pro with P and Thr, Tyr with T) but that initial letter is used as one-letter-amino-acid-code for only one of these (see Discussion and Codons and amino acids)
    • initiating methionine (Met1) - p.Met1? denotes that amino acid Methionine-1 (translation initiation site) is changed and that it is unclear what the consequence of this change is. When experimental data show that no protein is made, the description p.0 should be used. The description p.Met1Val is not allowed (see Discussion)
    • nonstop change - recommendations have recently been made to describe substitutions in the stop codon, so called nonstop changes like p.X110Tyrext16 (see Recommendations)
12. Polymorphisms - do not describe polymorphic variants as c.127A/G or p.43Ile/Val (or p.43I/V). A description of a variant should be neutral and polymorphisms and pathogenic changes should not be described differently (see Discussion). Correct descriptions are c.127A>G and p.Ile43Val.

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