Ann Lab Med.  2012 May;32(3):238-241. 10.3343/alm.2012.32.3.238.

Identification of a Rare 3 bp BRAF Gene Deletion in a Thyroid Nodule by Mutant Enrichment with 3'-Modified Oligonucleotides Polymerase Chain Reaction

• Jang MA
• Lee ST
• Oh YL
• Kim SW
• Chung JH
• Ki CS
• Kim JW

• KMID: 1381693
• DOI: http://doi.org/10.3343/alm.2012.32.3.238

Abstract

Papillary thyroid carcinoma (PTC) is the most common malignant thyroid tumor, and 36-69% of PTC cases are caused by mutations in the BRAF gene. The substitution of a valine for a glutamic acid (V600E) comprises up to 95-100% of BRAF mutations; therefore, most diagnostic methods, including allele-specific PCR and real-time PCR, are designed to detect this mutation. Nevertheless, other mutations can also comprise the genetic background of PTC. Recently, a novel and sensitive technique called mutant enrichment with 3'-modified oligonucleotides (MEMO) PCR has been introduced. When we applied allelespecific PCR and MEMO-PCR for the detection of the BRAF V600E mutation, we found an unusual 3' bp deletion mutation (c.1799_1801delTGA) only when using MEMO-PCR. This deletion results in the introduction of a glutamic acid into the B-Raf activation segment (p.V600_K601delinsE), leading to an elevated basal kinase activity of BRAF. This is the first report of a rare 3 bp BRAF deletion in a PTC patient that could not be detected by allele-specific PCR.

Keyword

Papillary thyroid carcinoma; BRAF; Deletion; Mutation; Mutant enrichment with 3'-modified oligonucleotides (MEMO) PCR; Korean

MeSH Terms

Alleles
Base Sequence
Female
Humans
Lymphatic Metastasis
Middle Aged
Oligonucleotides/genetics
Polymerase Chain Reaction/*methods
Proto-Oncogene Proteins B-raf/*genetics/metabolism
Sequence Deletion
Thyroid Neoplasms/genetics/pathology
Thyroid Nodule/metabolism

Figure

• 

  Fig. 1 Allele-specific PCR results. At left, two PCR products at 538 and 335 bp were amplified for the positive control of BRAF V600E; at right, the patient with the BRAF deletion mutation only showed a wild-type band at 538 bp.

• 

  Fig. 2 Direct sequencing of exon 15 of the BRAF gene after MEMO-PCR. Top, wild-type BRAF; bottom, a 3 bp deletion of coding nucleotides 1799 to 1801 (c.1799_1801delTGA; arrow). Abbreviation: MEMO-PCR, mutant enrichment with 3'-modified oligonucleotides-PCR.

• 

  Fig. 3 Confirmation of the deletion mutation using conventional PCR and sequencing. Top, wild-type BRAF; bottom, heterozygous peaks were observed from a starting position at nucleotide 1799 (arrow).

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