Tool For Molecular Diagnosis Research Articles

Cytogenetics and molecular genetics of carcinomas arising from thyroid epithelial follicular cells

Cytogenetic and molecular analyses of thyroid tumors have indicated that these neoplasms represent a good model for analyzing human epithelial cell multistep carcinogenesis. They comprise, in fact, a broad spectrum of lesions with different phenotypes and variable biological and clinical behavior. Molecular analysis has detected specific genetic alterations in the different types of thyroid tumors. In particular, the well-differentiated carcinomas of the papillary type are characterized by activation of the receptor tyrosine kinases (RTKs), RET and NTRKI proto-oncogenes. Cytogenetic analysis of these tumors has contributed to defining the chromosomal mechanisms leading to RTK oncogenic activation. In the majority of cases, intrachromosomal inversions of chromosome 10 and chromosome I led to the formation of RET-derived and NTRKI-derived oncogenes, respectively. Interestingly, molecular analysis of these oncogenes revealed their nature of chimeric fusion proteins all sharing the tyrosine kinase (TK) domains of the respective proto-oncogenes. Moreover, the sequencing of the oncogenic rearrangements led to the identification of a breakpoint cluster region in both RTK proto-oncogenes. Exposure to ionizing radiation is associated with papillary carcinomas and RET activation has been suggested to be related to this event. Conversely, RAS point mutations are frequently observed in tumor with follicular histology and have been associated with metastatic dissemination. Iodide-deficient areas seem to provide a higher-frequency of RAS positive follicular carcinomas. Finally, a high prevalence of TP53 point mutations has been detected only in undifferentiated or anaplastic carcinomas and found to correlate inversely with BCL2 expression. All of these findings are contributing to the definition of genetic and environmental factors relevant for the pathogenesis of thyroid tumors. Moreover, the characterization of specific genetic lesions could provide significant molecular tools for a better differential diagnosis and for the development of novel therapeutic avenues for thyroid cancer. Genes Chromosom Cancer 16:1–14 (1996). © 1996 Wiley-Liss, Inc.

Molecular tools for presymptomatic testing in multiple endocrine neoplasia type 1

The aim of this workshop session was to define a set of molecular tools for pre-clinical diagnosis in affected families, and to assess presence or absence of linkage to 11q13 in families with classical MEN1 as well as with MEN1-related clinical features. A consensus linkage map of first- and second-choice markers based on PCR as well as Southern blotting was established at the workshop. Based on the results from linkage analysis in 87 families with classical MEN1, presymptomatic testing using the suggested panel of markers, can now be performed with great accuracy.

Self-fusion of the ALL 1 Gene

LEUKEMIA is a clonal malignancy of blood cells that usually involves the myeloid or lymphoid lineages. The origin of leukemia, like other forms of cancer, lies in gene defects that alter the normal programs of cellular differentiation, proliferation, or apoptosis. Using chromosomal rearrangements as a guidepost to the location of genes involved in hematologic malignancy, researchers have identified and molecularly characterized many leukemia genes during the last decade. 1 These studies have revealed some common genetic mechanisms for leukemogenesis, provided molecular tools for leukemia diagnosis and monitoring, and identified potential targets for a new generation of leukemia therapies. One of the most significant of these genetic findings involves the ALL1 gene, which participates in acute leukemias in 5% to 10% of children and adults. Like other leukemia genes, ALL1 was discovered by its association with well-characterized chromosomal rearrangements. However, we have recently found that the ALL1 gene also participates in

Analysis of the OA1 gene reveals mutations in only one-third of patients with X-linked ocular albinism.

The locus for ocular albinism type 1 (OA1) has been assigned to the Xp22.3 region through both linkage and deletion mapping. The disorder was found to be genetically homogeneous, as all informative families showed convincing linkage data with markers on Xp22.3 and all identified deletions involved in the same region. The OA1 gene recently was cloned and several intragenic deletions were identified in affected individuals. We have characterized the genomic structure of the OA1 gene, which spans approximately 40 kb of genomic DNA and contains nine exons. A highly polymorphic dinucleotide repeat was identified in intron 1, that provides a useful tool for molecular diagnosis. Knowledge of the intron/exon boundaries allowed us to search for point mutations in patients' genomic DNA. All nine exons of the OA1 gene, as well as the 5' and 3' untranslated regions, were scanned for point mutations in PCR-amplified DNA from 60 OA1 patients. The mutations identified include: two frameshifts and a splice site mutation leading to truncated OA1 proteins; a deletion of a threonine codon at position 290; and four missense mutations,two of which involve amino acids located within putative transmembrane domains. Two of the mutations each occur in three apparently unrelated families, consistent with previous observations of a founder effect in OA1. Surprisingly, mutations were detected in only one-third of the patients (21 of 60) ascertained. We postulate that mutations not yet identified in either regulatory elements of the OA1 gene, or in other gene(s) located within the same chromosomal region, may be common cause of X-linked ocular albinism.

DNA fingerprinting for differentiating aggressivity groups of the rape seed pathogen Leptosphaeria maculans

Genomic DNA was prepared from a total of 16 different isolates of the rape seed pathogen Leptosphaeria maculans , the teleomorph of Phoma lingam . These DNA preparations were digested with the restriction endonucleases Bam H I, Eco R I, Hpa II, Mbo I, Msp I and Sau3 A I, separated on agarose gels and analysed by hybridization with the radioactively labelled synthetic oligonucleotides (GTG) 5 , (GACA) 4 or (CT) 8 or with labelled DNA of phage M13. These probes reveal a high degree of DNA polymorphism within the species. Although all investigated isolates can clearly and unequivocally be distinguished from each other, aggressive and nonaggressive strains appear as consistent groups with high degrees of similarity. The fingerprinting technique is thus an appropriate tool for molecular diagnosis of this important plant pathogen. In general, oligonucleotide fingerprinting may be regarded as a convenient technique to identify individual fungal strains with a reliability similar to that of mammalian systems.