Analysis of transgenerational effects on DNA copy number aberrations in male mice exposed to continuous 20mGy/day gamma-rays for 400 days (Primary screening for 0mGyG family). Mus musculus

Transgenerational effects of continuous low dose-rate (LDR) gamma-ray irradiation have not been well studied. Recent advances in DNA technology enabled us to examine a whole genome at molecular level. Here we adopted one of these techniques called oligo-microarray comparative genome hybridization (CGH) and studied trans-generational effects on DNA copy number aberrations (CNAs). C57BL/6JNrs male mice were exposed to LDR gamma-rays (20 mGy/day) for 400 days (total dose: 8000 mGy) from 8 weeks of age. Progeny from 20 mGy/day irradiated mice had significantly higher frequencies of genomic aberrations than the progeny of non-irradiated mice. This study investigated the de novo mutation by comparing the parents and children using 25 irradiated male family and 25 non-irradiated male family. Grant: This study was performed under contract with the Aomori Prefectural Government, Japan. Overall design: We irradiated and non-irradiated male mice (C57BL/6JNrs) for 400 days and crossed to non-irradiated female mice (C57BL/6JNrs) and got F1 mice. After they were dead, the DNA were prepared from their tails. At first, primary screening using 1 M oligo-microarray was performed. The array was mounted autosomal fragment at intervals of an average about 3kb. Two-condition experiment, mice tail tissue irradiated or non-irradiated male, female and their progenies vs. reference male mouse. This reference male mouse was from the same colony used in this experiment. Twice by changing the labeling color Cy3 and Cy5, performed CGH and extracting probes showed either aberration. Identification of the chromosomal location of the probe. Probes adjacent to the probe with aberration were set at high density. The candidate probes in one family was designed in a single 4x 244k or 8x 60k oligo-microarray and performed secondary screening. Extracting the flanking probes showed either aberration when labeled with Cy3 and Cy5 and identification the chromosomal location of the probe. Finally, we performed TaqMan® Copy Number Assays to verify the mutations. This SubSeries is a primary screening. Non-irradiated male (0mGyGM), female (0mGyGF) and their progenies (0mGyG1 to 0mGyG6) vs. reference male mouse 1.

持续低剂量率（low dose-rate, LDR）γ射线辐照的跨代效应尚未得到充分研究。DNA技术的最新进展使我们能够在分子层面对全基因组进行检测分析。本研究采用其中一项名为寡核苷酸微阵列比较基因组杂交（oligo-microarray comparative genome hybridization, CGH）的技术，探究了辐照对DNA拷贝数畸变（copy number aberrations, CNAs）的跨代影响。

C57BL/6JNrs雄性小鼠于8周龄起，暴露于低剂量率γ射线（20 mGy/天），持续400天，总辐照剂量为8000 mGy。辐照组小鼠的子代基因组畸变频率显著高于未辐照组小鼠的子代。本研究通过对比25个辐照雄性家系与25个未辐照雄性家系的亲子代样本，对新生突变进行了分析。

资助说明：本研究依托日本青森县政府的合同项目开展。

实验整体设计：将C57BL/6JNrs雄性小鼠分为辐照组与对照组，辐照组持续辐照400天，随后与未辐照的同品系雌性小鼠交配，获得F1代小鼠。待小鼠处死后，从其尾部组织提取基因组DNA。本研究首先采用1M寡核苷酸微阵列进行初步筛选。该微阵列针对常染色体片段设计，探针平均间隔约3kb。实验采用双条件分组：将辐照/未辐照的雌雄小鼠及其子代的尾部组织，与来源于本实验同一饲养群的参照雄性小鼠样本进行对比。通过交替使用Cy3与Cy5两种荧光标记染料，重复开展两次CGH实验，筛选出存在畸变信号的探针，并对其进行染色体定位；同时针对存在畸变的探针及其邻近区域设计高密度探针。针对单个家系的候选探针，将其设计在单张4×244k或8×60k寡核苷酸微阵列上，进行第二轮筛选：再次筛选经Cy3与Cy5标记后存在畸变信号的侧翼探针，并完成探针的染色体定位。最后，采用TaqMan®拷贝数检测试剂盒对突变位点进行验证。

本子系列为初步筛选数据集：将未辐照雄性小鼠（0mGyGM）、雌性小鼠（0mGyGF）及其子代（0mGyG1至0mGyG6）与参照雄性小鼠1号进行对比。