EUROPEAN RADIATION RESEARCH 2012

Estimation of transgenerational effects of radiation in male germ cells of mice using high-density microarray CGH platform

16 Oct 2012, 16:30

1m

Poster Hall (Vietri sul Mare)

poster preferred Other Poster Session 1

Speaker

Dr Jun-ichi Asakawa (Dept. of Genetics, Radiation Effects Research Foundation)

Description

We used two methods to estimate genetic effects of radiation. The first method was the comparative genomic hybridization (CGH) with high-density microarray containing 2.1 million probes (HD-2 microarray), which permits high-resolution examination of copy number changes (CNV: deletions and amplifications) throughout the whole genomes between two DNA samples. Refinement of the experimental and analytical protocols allowed accurate detection of small (2-3 kb, harboring only two adjacent probes) to large deletions which were previously characterized. The HD-2 microarray CGH approach was used for detection of deletion/amplification mutations among the genomes of 80 F1 mice derived from 4-Gy gamma irradiated spermatogonia and 80 control mice. A total of 22 mutations, 10 in the exposed group (6 deletions in 6 mice, 4 amplifications in 4 mice) and 12 mutations in the control (7 deletions in 7 mice, 5 amplifications in 3 mice, i.e., 1 mouse had 3 amplifications) were detected. The second was Restriction Landmark Genome Scanning (RLGS), a method which visualizes several thousand DNA fragments as spots and permits the detection of an autosomal deletion as a half-normal intensity spot. We applied the RLGS method for a genome-wide assessment of the induction rate of deletion mutations. Examinations were made on 1,007 progeny (502 control and 505 derived from spermatogonia exposed to 4 Gy of X rays). The results showed 1 deletion mutation in the un-irradiated paternal genomes of 502 offspring (0.2%) and 5 deletions in the irradiated paternal genomes of 505 offspring (1%). The deletion sizes ranged from 2 to 13 Mb. If the frequencies are taken at face values, the net increase was 4 deletions after an exposure of 4 Gy, or 1 deletion per Gy per individual if a linear dose response is assumed. Since the present RLGS screened about 0.2% of the total genome, the probability of any deletion mutation induced in the whole genome is estimated as 500 times (i.e. 1/0.002) of 0.2%, or about 1 per Gy. On the other hand, the CGH method using HD-2 microarray, containing 2.1 million probes distributed at approximately 1-kb intervals, searched deletion mutations at about 70% of the whole genome. The number of deletion mutation in the 80 exposed offspring is much smaller than that expected from the RLGS results (about 220; 80 × 1 deletion/Gy ×４Gy × 70%). The results imply that the number of large deletions induced by radiation exposure maybe considerably smaller than currently estimated.