School of Agriculture and Food Science, University College Dublin, Belfield, Dublin, Ireland.
PLoS One. 2024 May 31;19(5):e0294089. doi: 10.1371/journal.pone.0294089. eCollection 2024.
Hair follicles provide an easily accessible tissue for interrogating gene expression for multiple purposes in mammals. RNAlater® is a liquid storage solution that stabilises and preserves cellular RNA, eliminating the need to immediately process or freeze tissue specimens. The manufacturer advises storage of samples at 2-8°C overnight before transfer to -20°C. This study aimed to evaluate RNA integrity in hair follicle samples collected from horses, stabilized in RNAlater®, and stored under three short-term storage conditions. Mane hair samples complete with follicles were collected from four horses at a single time point. Approximately 15 hairs were placed in each of three 2 mL tubes containing 0.75ml RNAlater® solution. Test group A was stored at 4°C for 24-h, then decanted and stored at -20°C. Test groups B and C were stored at 4°C and 19°C (room temperature) respectively for 7 days, then decanted and stored at -20°C. RNA was isolated from all samples and RNA quantity and quality were measured. One-way ANOVA revealed no difference in RNA concentration (A:516 +/-125 ng/ml, B:273+/-93 ng/ml, C:476+/-176 ng/ml;P = 0.2) or quality (A:9.5 +/-0.19, B:9.8+/-0.09, C:9.2+/-0.35 RIN; P = 0.46) between the test groups. There were no group differences in mean Cycle Threshold values from qPCR validation assays confirming high-quality template cDNA. The results suggest that storage of hair follicles for one week in RNAlater® at cool or room temperature conditions will not compromise RNA integrity and will permit extended transport times from remote sampling locations without the need for freezing.
毛囊为哺乳动物提供了一种易于获取的组织，可用于多种目的的基因表达分析。RNAlater®是一种液体储存溶液，可稳定和保存细胞 RNA，从而无需立即处理或冷冻组织标本。制造商建议将样本在 2-8°C 下储存过夜，然后再转移至-20°C。本研究旨在评估 RNAlater®稳定的马毛囊样本的 RNA 完整性，并在三种短期储存条件下储存。在一个时间点从四匹马的马鬃毛中收集完整的毛囊样本。每个 2 mL 管中放入大约 15 根毛发，管中含有 0.75ml RNAlater®溶液。A 组在 4°C 下储存 24 小时，然后倒出并储存在-20°C。B 组和 C 组分别在 4°C 和 19°C（室温）下储存 7 天，然后倒出并储存在-20°C。从所有样本中提取 RNA，并测量 RNA 数量和质量。单因素方差分析显示，RNA 浓度（A：516+/-125ng/ml，B：273+/-93ng/ml，C：476+/-176ng/ml；P=0.2）或质量（A：9.5+/-0.19，B：9.8+/-0.09，C：9.2+/-0.35 RIN；P=0.46）在实验组之间没有差异。qPCR 验证实验的平均循环阈值没有组间差异，证实了高质量的模板 cDNA。结果表明，将毛囊在 RNAlater®中在凉爽或室温条件下储存一周不会影响 RNA 完整性，并允许从偏远采样地点延长运输时间，而无需冷冻。
