An Effi cient Method for Total RNA Extraction of Poplar Bark Infected with Pathogen and the Application
Аннотация
The highly efficient total RNA extraction of poplar bark infected with pathogen is important for the expression and regulation research of resistance genes. To explore an efficient method for total RNA extraction from the lesion bark, 2-year-old seedling stem of Populus euramericana cv. ‘Zhonglin 46' was inoculated with the pathogenic bacterium of Lonsdalea quercina subsp. populi and used for RNA extraction. A novel RNA extraction method was first proposed in this study and named Improved method of RNA extraction kit. We compared the extraction effects among this method and other five methods. The test results showed that RNA extracted using Improved method of RNA extraction kit was of high concentration and the band of 28S rRNA was nearly twice as bright as that of 18S rRNA. The analysis suggested that Improved method of RNA extraction kit was the most suitable for total RNA extraction from the lesion bark. To test the wider applicability, we further extracted total RNA from other tissues, including the healthy bark of P. euramericana cv. ‘Zhonglin 46', P. euramericana cv. ‘74/76' and Populus×beijingensis, tissue culture plantlets of Populus tomentosa treated with low nitrogen or phosphorus stress and the fl ower of Magnolia denudate. The testing showed that total RNA could be successfully extracted from all these plant tissues with high quality. The extracted RNA had been successfully used for transcriptome sequencing of poplar lesion bark, RT-PCR and qRT-PCR of tissue culture plantlets of P. tomentosa treated with low nitrogen stress. Consequently, the result showed that total RNA extracted using Improved method of RNA extraction kit could be further used in the subsequent molecular experiment.