The main intent of northern blotting is an essential strategy in molecular biology to quantify the particular messenger RNA (mRNA). The very initial step is to identify the cells that express a specific gene, therefore providing a better understanding regarding the physiological role of the protein that has been encoded. Understanding the nutritional, hormonal, or environmental factors which governed by a particular gene is the primary aim of measuring mRNA. In addition to identifying transcript presence, it also provides size and a comparative analysis for transcript abundance over the single membrane. It’s been effectively adapted in recent years to investigate and validate the expression in addition to size of short noncoding RNA. The denaturing agarose/polyacrylamide gel should be used to segregate RNA based on their size before it can be transferred on the solid surface in a way that maintains the topology distribution of the gel for the purpose of preparing northern blot to hybridization. This chapter clearly emphasizes the step-by-step protocol for northern blotting.