The first one is the annealing temperature (annealing temperature is the temperature parameter when primers and templates are combined). When 50% of the primers and complementary sequences are expressed as double-stranded molecules, they are important factors affecting the specificity of PCR. Ideally, the annealing temperature is low enough to ensure that the primers are annealed effectively to the desired sequence, while still high enough to reduce non-specific binding. The reasonable annealing temperature ranges from 55 C to 70 C. The annealing temperature is usually set to be 5 C lower than the Tm of the primer.


Does it reach the specified temperature? Large-scale flange treatment is usually carried out by solution heat treatment, commonly known as "annealing", and the temperature range is 1040-1120 ("Japan Standard"). You can also observe through the observation hole of the annealing furnace. Large flanged joints in annealing zone should be incandescent, but without softening and sagging.


Secondly, the protective gas pressure, in order to prevent the occurrence of micro-leakage, the protective gas in the furnace should maintain a certain positive pressure. If it is hydrogen gas, it generally needs more than 20 kBar. Annealing atmosphere: Usually pure hydrogen is used as annealing atmosphere, and the purity of the atmosphere is preferred to be 99.99% or higher. If the other part of the atmosphere is inert gas, the purity can be lower, but it should not contain too much oxygen or water vapor.


Another point is the sealing of the furnace (the shell is welded by steel plate and profiled steel, the car is welded by steel plate and steel plate, and the car reduces heat radiation and convection loss through soft contact with the lining and sand sealing mechanism). The bright annealing furnace should be isolated from the outside air; hydrogen is used as a protective gas, and only one outlet is opened.（ Used to ignite the exhausted hydrogen. The inspection method can be applied to the gap of annealing furnace joint with soap water to check whether the gas is in operation. The easiest place to run gas is where the annealing furnace enters the pipeline and where the pipeline discharges. The sealing rings in this place are particularly easy to wear out. Check regularly for changes.