A New Biotin Labeling and High-Molecular-Weight RNA Northern Method and Its Application in Viral RNA Detection
Xi Zhang,
Qingling Zhang,
Long Cheng,
Dan Liu,
Hongzheng Wang,
Yingjia Zhou,
Liqun Ma,
Jubin Wang,
Feng Li
Affiliations
Xi Zhang
Key Laboratory of Horticultural Plant Biology (MOE), College of Horticulture and Forestry Sciences, Huazhong Agricultural University, Wuhan 430070, China
Qingling Zhang
Key Laboratory of Horticultural Plant Biology (MOE), College of Horticulture and Forestry Sciences, Huazhong Agricultural University, Wuhan 430070, China
Long Cheng
Key Laboratory of Horticultural Plant Biology (MOE), College of Horticulture and Forestry Sciences, Huazhong Agricultural University, Wuhan 430070, China
Dan Liu
Key Laboratory of Horticultural Plant Biology (MOE), College of Horticulture and Forestry Sciences, Huazhong Agricultural University, Wuhan 430070, China
Hongzheng Wang
Key Laboratory of Horticultural Plant Biology (MOE), College of Horticulture and Forestry Sciences, Huazhong Agricultural University, Wuhan 430070, China
Yingjia Zhou
Key Laboratory of Horticultural Plant Biology (MOE), College of Horticulture and Forestry Sciences, Huazhong Agricultural University, Wuhan 430070, China
Liqun Ma
Key Laboratory of Horticultural Plant Biology (MOE), College of Horticulture and Forestry Sciences, Huazhong Agricultural University, Wuhan 430070, China
Jubin Wang
Key Laboratory of Horticultural Plant Biology (MOE), College of Horticulture and Forestry Sciences, Huazhong Agricultural University, Wuhan 430070, China
Feng Li
Key Laboratory of Horticultural Plant Biology (MOE), College of Horticulture and Forestry Sciences, Huazhong Agricultural University, Wuhan 430070, China
Viruses cause severe crop losses. Studying the interaction between viruses and plants is very important for development of control measures. Northern blot is a well-accepted but very challenging technique to monitor the infection of viruses. Here, we modified the high-molecular-weight (hmw)RNA Northern blot experiment process, utilizing vertical electrophoresis to separate the RNA with denatured agarose gel. This protocol is compatible with regular equipment for Western blots and small RNA Northern blots and requires less input of total RNA. A new method to label the probe with biotin was also developed, which requires commonly used T4 DNA polymerase and detects viral RNA with high sensitivity. These new protocols made hmwRNA Northern blot cost-effective and easy-to-operate, very suitable for studying virus–host interactions.
