Preparation of culture medium

The mother liquor is generally pre-prepared into a mother liquor which is 10 times higher than the required concentration, which is usually stored in the refrigerator and only needs to be taken according to the proportion when preparing the culture medium.

Taking MS medium as an example, according to the characteristics of various drugs, the mother liquor can be divided into 6 kinds (see table).

┌───┬─┬─┐

│ │ to prepare │ quantity │

├───┼─┼─┤

When 1 │ NHlNO3 82.5g │ 50 solution of │ mother liquor was mixed with 1 liter medium, 20ml │ was obtained.

│ │ KNO3 950g │ │

│ │ MgSO4. 7H2O 18. 5g │ │

│ │ add water to fix volume to 1000 ml │ │

├───┼─┼─┤

When │ mother liquor 2 │ CaCI2 22.0g │ 100x solution was mixed with 1 liter medium, 10ml │ was taken.

│ │ add water to 500ml │ │

├───┼─┼─┤

When │ mother liquor 3 │ KH2PO4 8. 5g │ 100x solution and 1 liter culture medium, take 10ml │

│ │ add water to 500ml │ │

├───┼─┼─┤

When 4 │ Na2-EDTA 3.73g │ 1DO doubling solution of │ mother liquor was mixed with 1 liter medium, 10ml │ was obtained.

│ │ FeSO4 7H2O 2.78g │ │

│ │ add water to fix volume to 1000 ml │ │

├───┼─┼─┤

│ mother liquor 5 │ H3BO3 620mg │ │

│ │ MnSO4 4H2O 2230 mg │ 100x solution with 1 liter medium, take 10 ml │

│ │ ZnSO4 7H2O 860mg │ │

│ │ KL 83 mg │ │

│ │ Na2MoO4 ZH2O 12.5mg │ │

│ │ CuSO4 ·5H2o 1.25mg │ │

│ │ CoCl2 ·6H2O 1.25mg │ │

│ │ add water to fix volume to 1000 ml │ │

├───┼─┼─┤

When │ mother liquor 6 │ inositol 5g │ 100x solution mixed with one liter medium, take 10ml │

│ │ glycine 100mg │ │

│ │ nicotinic acid 25 mg │ │

│ │ 25 mg │ │ Hydrochloride

│ │ 5mg │ │ Hydrochloride

│ │ add water to 500ml │ │

└───┴─┴─┘

The prepared mother liquor bottle should be labeled with the mother liquid number, the preparation multiple, the date and the amount that should be taken when mixing 1 liter of mother liquid. The prepared mother liquor can be stored in the refrigerator (it can be preserved at low temperature for several months). If mold and precipitation are found, it can not be used.

When preparing the culture medium, we should first make various preparations: first, arrange the mother liquor in order, then prepare all kinds of glassware needed, weigh the necessary Agar swelling and sucrose, and match the necessary growth regulators.

When preparing, first put a certain amount of ice in a simple or other form of container to avoid precipitation caused by splashing or mixing with too high concentration. Then according to the order of the mother liquid, according to the prescribed quantity. According to the need to add some of the most growth regulatory substances, and finally add Agar and sucrose, after heating and melting in the pot. The prepared medium should be adjusted by pH with D.L ~ and equivalent HCI (or NaOH), which is generally kept in the range of 5.1mur6.0. The prepared culture medium should be injected separately while it is hot and put into the incubator such as test tube and flask.

After the culture medium is equipped, it should be sterilized under high pressure in time, and it can be inoculated and prepared after cooling and condensation.

3. The use of growth regulators

Growth regulators are the key substances in the culture medium, which play a decisive role in plant tissue culture.

The main results are as follows: (1), auxins include NAA (Cai acetic acid), IAA (indoleacetic acid), IBA (indolebutyric acid) and so on. It is very sensitive to the growth of roots, followed by the growth of buds and stems. It is the main promoting substance in rooting culture of tissue culture.

(2) the cytokinins include BA (6-phenylpurine), KT (kinetin), ZT (zeatin) and so on. It can promote cell division and bud growth. It binds with a small amount of auxin and often induces the proliferation and growth of buds.

(3) Natural extract. Some natural substances can promote the growth of plant tissues and cells. (Wang Huaiyu)