Martina Auer

Publications of Martina Auer

• Combined molecular genetic and cytogenetic analysis from single cells after isothermal whole-genome amplification.

  Authors: Thomas Kroneis, Jochen B Geigl, Amin El-Heliebi, Martina Auer, Peter Ulz, Thomas Schwarzbraun, Gottfried Dohr, Peter Sedlmayr

  Clinical chemistry. 05/2011; 57(7):1032-41.

  Analysis of chromosomal aberrations or single-gene disorders from rare fetal cells circulating in the blood of pregnant women requires verification of the cells' genomic identity. We have developed aAnalysis of chromosomal aberrations or single-gene disorders from rare fetal cells circulating in the blood of pregnant women requires verification of the cells' genomic identity. We have developed a method enabling multiple analyses at the single-cell level that combines verification of the genomic identity of microchimeric cells with molecular genetic and cytogenetic diagnosis. We used a model system of peripheral blood mononuclear cells spiked with a colon adenocarcinoma cell line and immunofluorescence staining for cytokeratin in combination with DNA staining with the nuclear dye TO-PRO-3 in a preliminary study to define candidate microchimeric (tumor) cells in Cytospin preparations. After laser microdissection, we performed low-volume on-chip isothermal whole-genome amplification (iWGA) of single and pooled cells. DNA fingerprint analysis of iWGA aliquots permitted successful identification of all analyzed candidate microchimeric cell preparations (6 samples of pooled cells, 7 samples of single cells). Sequencing of 3 single-nucleotide polymorphisms was successful at the single-cell level for 20 of 32 allelic loci. Metaphase comparative genomic hybridization (mCGH) with iWGA products of single cells showed the gains and losses known to be present in the genomic DNA of the target cells. This method may be instrumental in cell-based noninvasive prenatal diagnosis. Furthermore, the possibility to perform mCGH with amplified DNA from single cells offers a perspective for the analysis of nonmicrochimeric rare cells exhibiting genomic alterations, such as circulating tumor cells.

• Evolution of genomic instability in diethylnitrosamine-induced hepatocarcinogenesis in mice.

  Authors: Kristina Aleksic, Carolin Lackner, Jochen B Geigl, Martina Schwarz, Martina Auer, Peter Ulz, Maria Fischer, Zlatko Trajanoski, Marcus Otte, Michael R Speicher

  Hepatology (Baltimore, Md.). 03/2011; 53(3):895-904.

  Diethylnitrosamine (DEN) is a hepatic procarcinogen which is frequently used as an inducer of hepatocellular carcinoma (HCC) in mice. Although mice after DEN exposure are among the most widely usedDiethylnitrosamine (DEN) is a hepatic procarcinogen which is frequently used as an inducer of hepatocellular carcinoma (HCC) in mice. Although mice after DEN exposure are among the most widely used models for liver tumorigenesis, a detailed, mechanistic characterization of the longitudinal changes in the respective tumor genomes has never been performed. Here we established the chronological order of genetic alterations during DEN carcinogenesis by examining mice at different points in time. Tumor samples were isolated by laser microdissection and subjected to array-comparative genomic hybridization (array-CGH) and sequencing analysis. Chromosomal gains and losses were observed in tumors by week 32 and increased significantly by week 56. Loss of distal chromosome 4q, including the tumor suppressors Runx3 and Nr0b2/Shp, was a frequent early event and persisted during all tumor stages. Surprisingly, sequencing revealed that β-catenin mutations occurred late and were clearly preceded by chromosomal instability. Thus, contrary to common belief, β-catenin mutations and activation of the Wnt/β-catenin pathway are not involved in tumor initiation in this model of chemical hepatocarcinogenesis. CONCLUSION: Our study suggests that the majority of the current knowledge about genomic changes in HCC is based on advanced tumor lesions and that systematic analyses of the chronologic order including early lesions may reveal new, unexpected findings.

• Mapping of balanced chromosome translocation breakpoints to the basepair level from microdissected chromosomes.

  Authors: Anna C Obenauf, Thomas Schwarzbraun, Martina Auer, Eva M Hoffmann, Julie Waldispuehl-Geigl, Peter Ulz, Barbara Günther, Hans-Christoph Duba, Michael R Speicher, Jochen B Geigl

  Journal of cellular and molecular medicine. 08/2010; 14(8):2078-84.

  The analysis of structural variants associated with specific phenotypic features is promising for the elucidation of the function of involved genes. There is, however, at present no approach allowingThe analysis of structural variants associated with specific phenotypic features is promising for the elucidation of the function of involved genes. There is, however, at present no approach allowing the rapid mapping of chromosomal translocation breakpoints to the basepair level from a single chromosome. Here we demonstrate that we have advanced both the microdissection and the subsequent unbiased amplification to an extent that breakpoint mapping to the basepair level has become possible. As a case in point we analysed the two breakpoints of a t(7;13) translocation observed in a patient with split hand/foot malformation (SHFM1). The amplification products of the der(7) and of the der(13) were hybridized to custom-made arrays, enabling us to define primers at flanking breakpoint regions and thus to fine-map the breakpoints to the basepair level. Consequently, our results will also contribute to a further delineation of causative mechanisms underlying SHFM1 which are currently unknown.

• Identification of small gains and losses in single cells after whole genome amplification on tiling oligo arrays.

  Authors: Jochen B Geigl, Anna C Obenauf, Julie Waldispuehl-Geigl, Eva M Hoffmann, Martina Auer, Martina Hörmann, Maria Fischer, Zlatko Trajanoski, Michael A Schenk, Lars O Baumbusch, Michael R Speicher

  Nucleic acids research. 07/2009;

  Clinical DNA is often available in limited quantities requiring whole-genome amplification for subsequent genome-wide assessment of copy-number variation (CNV) by array-CGH. In pre-implantationClinical DNA is often available in limited quantities requiring whole-genome amplification for subsequent genome-wide assessment of copy-number variation (CNV) by array-CGH. In pre-implantation diagnosis and analysis of micrometastases, even merely single cells are available for analysis. However, procedures allowing high-resolution analyses of CNVs from single cells well below resolution limits of conventional cytogenetics are lacking. Here, we applied amplification products of single cells and of cell pools (5 or 10 cells) from patients with developmental delay, cancer cell lines and polar bodies to various oligo tiling array platforms with a median probe spacing as high as 65 bp. Our high-resolution analyses reveal that the low amounts of template DNA do not result in a completely unbiased whole genome amplification but that stochastic amplification artifacts, which become more obvious on array platforms with tiling path resolution, cause significant noise. We implemented a new evaluation algorithm specifically for the identification of small gains and losses in such very noisy ratio profiles. Our data suggest that when assessed with sufficiently sensitive methods high-resolution oligo-arrays allow a reliable identification of CNVs as small as 500 kb in cell pools (5 or 10 cells), and of 2.6-3.0 Mb in single cells.