由于多糖的复杂支架,从成熟的椰子组织中获取高级RNA是一个棘手且劳动密集型的过程,多酚,脂质,以及与核酸形成牢固复合物的蛋白质。然而,我们第一次有效地开发了一种新的方法,从根部提取高级RNA,胚乳,和成熟椰子树的中果皮发生。在这种方法中,我们利用二氯甲烷代替苯酚/氯仿,从成熟的椰子组织中回收RNA。从成熟椰子树的根部获得的高级RNA的量为120.7µg/g,A260/280比率为1.95。同样,成熟的椰子中果皮产生134.6µg/gFW的优质RNA,A260/280比率为1.98,而成熟的椰子胚乳产生120.4µg/gFW的优质RNA,A260/280比率为2.01。此外,使用二氯甲烷方法的RNA分离在下游实验中表现出优异的性能,特别是在用于cDNA生产和扩增的RT-PCR中。相反,RNA植物试剂盒,TRIZOL,和十六烷基三甲基溴化铵(CTAB)方法在从上述椰子组织中分离出大量纯度异常的RNA方面均未成功。鉴于这些发现,我们得出的结论是,新开发的方法将在有效地从成熟的椰子组织中提取高纯度的RNA方面至关重要。
Procuring high-grade RNA from mature coconut tissues is a tricky and labor-intensive process due to the intricate scaffold of polysaccharides, polyphenols, lipids, and proteins that form firm complexes with nucleic acids. However, we have effectively developed a novel method for the first time, letting the retrieval of high-grade RNA from the roots, endosperm, and mesocarp of mature coconut trees take place. In this method, we exploited
dichloromethane as a replacement to phenol/chloroform for RNA recovery from mature coconut tissues. The amount of high-grade RNA acquired from the roots of mature coconut trees was 120.7 µg/g, with an A260/280 ratio of 1.95. Similarly, the mature coconut mesocarp yielded 134.6 µg/g FW of quality RNA with A260/280 ratio of 1.98, whereas the mature coconut endosperm produced 120.4 µg/g FW of quality RNA with A260/280 ratio of 2.01. Furthermore, the RNA isolation using the
dichloromethane method exhibited excellent performance in downstream experiments, particularly in RT-PCR for cDNA production and amplification. On the contrary, the RNA plant kit, TRIZOL, and Cetyl Trimethyl Ammonium Bromide (CTAB) methods were unsuccessful in isolating substantial quantities of RNA with exceptional purities from the mentioned coconut tissues. In view of these findings, we conclude that the newly developed method will be pivotal in effectively extracting RNA with high purity from mature coconut tissues.