The success and reliability of experiments in a plant tissue culture laboratory depend upon the design of the laboratory layout and the maintenance of sterile conditions. This chapter outlines essential aspects of laboratory organization, including the design of sterile zones, availability of equipment, workflow management, record-keeping, and quality control measures. Maintaining sterility is crucial for experimental success, with practices such as aseptic manipulations, air quality monitoring, microbial testing via surface swabbing, and visual inspections being vital for preventing contamination. Adherence to standard operating procedures (SOPs) for media preparation, culture handling, equipment sterilization, cleaning, waste disposal, and emergency response ensures consistency and compliance with sterile techniques. A well-designed laboratory layout, strict sterile practices, and best laboratory practices are key for successful tissue culture, propagation, and secondary metabolite production. The culture medium is critical, providing essential nutrients, growth regulators, and a controlled environment for cell growth and development. It influences differentiation, organogenesis, and secondary metabolite production. An optimal medium supports growth, minimizes contamination, and enhances nutrient uptake. This chapter serves as a guide for researchers aiming to establish a facility for secondary metabolite production, general techniques practiced in plant tissue culture laboratories.