An improved method for isolation of RNA from bone

We developed a simple approach to isolate bone RNA that combines pulverizing the bone and the phenol-guanidinium based RNA extraction in a single step while maintaining near-freezing temperatures. This single step method increases the yield of high quality RNA by eight-fold, with RNA integrity numbers ranging from 6.7 to 9.2.Our streamlined approach substantially increases the yield of high-quality RNA from bone tissue while facilitating safe and efficient processing of multiple samples using readily available platforms. The RNA obtained from this method is suitable for use in gene expression analysis in real-time quantitative PCR, microarray, and next generation sequencing applications.Obtaining intact, high quality RNA is an essential step in analyzing gene expression. This step is particularly challenging in bone, which contains low numbers of cells embedded within a highly mineralized tissue. As the endocrine functions of bone [1] and the relationship between bone and adipose physiology [2] becomes increasingly apparent, the need to isolate high quality RNA for gene expression analysis in bone using the current genome-wide sequencing technologies will gain more importance.Current methods for isolating RNA from bone use multiple steps in which the frozen bone is wrapped in foil, refrozen in liquid nitrogen and ground into a powder using a hammer [3] or ground using a mortar and pestle containing liquid nitrogen[4-6]. The powdered bone is then transferred to a second container for extraction of the RNA using a phenol-guanidinium-based reagent. While these approaches support extraction of RNA from bone, the multiple steps introduce the possibility of sample loss and the potential for cross-contamination. In addition, this approach does not lend itself to efficiently processing multiple samples. Herein, we report a one-step method for extracting RNA from bone (Figure 1) that consistently yields high-quality RNA suitable for gene expression analysis using the current