Detection of intronic polymorphism in heterozygous ductal breast carcinoma samples by non-isotopic RNase Cleavage Assay

Nordin N¹, Thilakavathy K¹, Seong NK², Sabariah AR² and Rosli R¹

1. Department of Human Growth and Development

2. Department of Clinical Laboratory Sciences, Faculty of Medicine and Health Sciences, Universiti Putra Malaysia, 43400 UPM, Serdang, Selangor, Malaysia.

Correspondence: Assoc Prof Dr Rozita Rosli; e-mail: This email address is being protected from spambots. You need JavaScript enabled to view it.

CITATION: Nordin N, Thilakavathy K, Seong NK, Sabariah AR, Rosli R. Detection of intronic polymorphism in heterozygous ductal breast carcinoma samples by non-isotopic RNase Cleavage Assay. International Medical Research Journal. 2002;6(1):33–40. 

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ABSTRACT

Alterations of p53 gene, a tumour-suppressor gene in various human malignancies, have been well characterised in half of human cancers, with I 0-30% alterations in breast carcinoma cases. An established screening method is important in order to reveal the p53 mutation patterns to strengthen its value as a predictive and prognostic marker in breast cancer. However, the wide variation in quality and quantity of the genomic DNA derived from clinical samples has often challenged the mutation detection strategies. Therefore, a reliable and sensitive detection method is needed in order to overcome the problem as well as to detect the various types of mutations scattered over the coding exons, especially exons 5-8, and their flanking intron sequences. The present study has managed to establish a mutation screening method using a non-isotopic RNase Cleavage Assay (NIRCA ™) from thirty-one invasive ductal breast carcinoma samples for intron 4 through exon 9 of the p53 gene. DNA sequence analysis was conducted to confirm the mutation spots screened by NIRCA ™. Interestingly, all samples studied showed intronic alterations with no mutation detected in the mutation hotspots of exons 5-8. Two intronic polymorphisms, as described elsewhere, were robustly detected in almost 50% of the samples screened by this method. This finding is important to our understanding of the biology of breast cancer and is an important contribution to the establishment of a Malaysian p53 mutation database.

KEYWORDS: Intronic mutation, p53, invasive ductal breast carcinoma, heterozygosity