CN104642132A

CN104642132A - Large-scale manufacturing method for high-precision AA culture medium

Info

Publication number: CN104642132A
Application number: CN201510082264.0A
Authority: CN
Inventors: 钱家静
Original assignee: Individual
Current assignee: Individual
Priority date: 2015-02-16
Filing date: 2015-02-16
Publication date: 2015-05-27

Abstract

The invention discloses a large-scale manufacturing method for a high-precision AA culture medium, and belongs to the technical field of plant biology. The large-scale manufacturing method comprises the following steps: firstly, calculating required amount, which is increased by 1000 times, of 16 elements contained in the medium as well as the required amount, which is increased by 10000 times, of CuSO4.5H2O and CoCl2.6H2O which have low required amount; accurately weighing the drugs; mixing and the 16 elements according to the required amount and sealing; dissolving CuSO4.5H2O and CoCl2.6H2O with 1L of distilled water for later use; and respectively opening according to the needs. The substrate prepared by the large-scale manufacturing method disclosed by the invention has the characteristics of being high in precision, convenient and quick to operate, simple to apply, and small in errors of different batches and within the batches, phenomena of large errors, precipitation, mildewing and the like caused by manual operation and external factor interferences during the preparation of high-precision AA liquid culture medium are avoided. The large-scale manufacturing method can be applied to the biotechnical purposes of plant tissue culture and the like.

Description

A kind of scale preparation method of high-precision A A culture matrix

Technical field

The invention discloses a kind of scale preparation method of high-precision A A culture matrix, belong to plant biotechnology field.Be more particularly one include accurate calculating, accurate weighing, first few after how mix, seal, mother liquor preparation separately, six steps such as use of breaking seal, to acquisition different batches and batch between the minimum culture matrix of error, minimizing experimental error method.

Background technology

Plant tissue culture technique or cultivation technique without soil need preparation medium mother liquor usually, and namely preparation expands certain multiple, liquid containing all kinds reagent.But, because mother liquor contains dissimilar reagent, be usually divided into the large class of macroelement, trace element, molysite and organic element four, and the multiple that four large classes expand is different, causes medicine service property (quality) inconsistent, easily occurs the error of calculation first; Second the producer of medicine, quality are inconsistent, even if to calculate, weigh consistent, also can affect result of the test; Three due to mother liquor be liquid condition, at the end of spring and the beginning of summer season, because the temperature rises and humidity increases, extremely easily cause mother liquor bacterial infection, mould, cause the phenomenon presenting different colours in mother liquor or occur precipitation, waste raw material; Comparatively big error can be there is to the difference of the cognition of container scale (especially trace element), instrument precision in four fundamental rules because of weighing person.

In view of the above-mentioned shortcoming of mother liquor; with the mother liquor medium of routine; usually there will be the failed phenomenon that maybe cannot repeat of the result of the test caused because of factors such as weighing person, medicine, seasons; have impact on the process using Plant Tissue Breeding or cultivation technique without soil to carry out test greatly, hit the confidence of experimenter.

Summary of the invention

A scale preparation method for high-precision A A culture matrix, is characterized in that, include in described method accurate calculating, accurate weighing, first few after how mix, seal, prepared by independent mother liquor, six steps such as use of breaking seal.

Method according to claim 1, is characterized in that, six described step concrete operations are as follows:

1. accurately calculate: 16 kinds of elements contained by AA minimal medium are expanded 1000 times, i.e. calcium chloride dihydrate 440g, potassium dihydrogen phosphate 170g, epsom salt 370g, potassium chloride 2940g, potassium iodide 0.83g, boric acid 6.2g, Sodium Molybdate Dihydrate 0.25g, white vitriol 8.6g, four water manganese sulphate 22.3g, iron edetate 36.5g, inositol 100g, thiamine hydrochloride 0.5g, glycine 75g, Glu 877g, L-Aspartic acid 266g, L-arginine 228g, 2 kinds of element CoCL2 6H2Os and cupric sulfate pentahydrate expand 10000 times, i.e. each 0.25g,

2. accurate weighing: according to the result precise of 1. middle calculating, especially with CoCL2 6H2O and the cupric sulfate pentahydrate of each 0.25g of ten thousand/balance precise;

3. how to mix after first few: mix according to few priority of carrying out of amount more, namely mix according to the order of Sodium Molybdate Dihydrate, thiamine hydrochloride, potassium iodide, boric acid, white vitriol, four water manganese sulphates, iron edetate, glycine, inositol, potassium dihydrogen phosphate, L-arginine, L-Aspartic acid, epsom salt, calcium chloride dihydrate, Glu, potassium chloride, amount to 5540.73g;

4. seal: the requirement of mixed solid matrix according to 250g every bottle is sub-packed in plastic containers, and seals;

5. mother liquor preparation separately: CoCL2 6H2O and cupric sulfate pentahydrate respectively weigh 0.25g, are dissolved in 1L distilled water, are mixed with 10000 times of mother liquors;

6. to break seal use: as required, add the above-mentioned solid matrix 5.54g of precise in every 1L distilled water, heating adds the independent mother liquor of 100 μ L CoCL2 6H2Os and cupric sulfate pentahydrate after it dissolves completely, and then adds reagent needed for other.

Beneficial effect of the present invention: the scale preparation method of this high precision culture matrix has following characteristics, i.e. without the error of calculation between the matrix of different batches; Because 16 kinds of reagent all expand 1000 times, 2 kinds of reagent expand 10000 times, and the weighing error between the matrix of different batches is minimum, and precision is high; In theory, due to the usage amount (calculating with each 1L) that solid matrix is 1000 times at every turn, namely the above-mentioned formula of 1000L can be prepared, liquid mother liquor only needs 100 μ L at every turn, and batch inside there will not be the human error caused because using the reagent of different manufacturers, different times; Solid matrix after expansion is that every 250g is sealed in plastic containers, can avoid because the impact of the extraneous factor such as weather, temperature, moisture causes occurring the common precipitation of liquid mother liquor, the phenomenon such as mouldy, and fluid matrix is only the mixing of two kinds of elements.After expanding 10000 times, precision is high; Above-mentioned plurality of advantages is all the follow-up test carried out for matrix with this medium, obtains result of the test accurately and lays the foundation.。

Embodiment:

Below in conjunction with specific embodiment, the present invention is conducted further description.

The scale preparation method of a kind of high-precision A A (carbonaceous sources and mounting medium) solid culture matrix in the present embodiment, include accurate calculating, accurate weighing, first few after how mix, seal, prepared by independent mother liquor, six steps such as use of breaking seal.

Above-mentioned six step concrete operations are as follows:

1. accurately calculate: 16 kinds of elements contained by AA medium are expanded 1000 times, i.e. calcium chloride dihydrate 440g, potassium dihydrogen phosphate 170g, epsom salt 370g, potassium chloride 2940g, potassium iodide 0.83g, boric acid 6.2g, Sodium Molybdate Dihydrate 0.25g, white vitriol 8.6g, four water manganese sulphate 22.3g, CoCL2 6H2O and each 0.025g of cupric sulfate pentahydrate, iron edetate 36.5g, inositol 100g, thiamine hydrochloride 0.5g, glycine 75g, Glu 877g, L-Aspartic acid 266g, L-arginine 228g;

2. accurate weighing: according to the result precise of 1. middle calculating, especially with CoCL2 6H2O and the cupric sulfate pentahydrate of each 0.025g of ten thousand/balance precise;

3. how to mix after first few: mix according to few priority of carrying out of amount more, namely mix according to the order of CoCL2 6H2O, cupric sulfate pentahydrate, Sodium Molybdate Dihydrate, thiamine hydrochloride, potassium iodide, boric acid, white vitriol, four water manganese sulphates, iron edetate, glycine, inositol, potassium dihydrogen phosphate, L-arginine, L-Aspartic acid, epsom salt, calcium chloride dihydrate, Glu, potassium chloride, amount to 5540.73g;

6. to break seal use: as required, add the above-mentioned matrix 5.54g of precise in every 1L distilled water, heating adds the independent mother liquor of 100 μ L CoCL2 6H2Os and cupric sulfate pentahydrate after it dissolves completely.

1L solution after above-mentioned substrate preparation is added the KOH of the 1N of 1ml, be sub-packed in the vial of 30 200ml, after 121 DEG C of sterilizing 20min.On superclean bench, inoculate photinia glabra callus, after cultivating 30d under 25 DEG C of dark conditions, callus disperses and well-grown, in milk yellow.

Claims

1. a scale preparation method for high-precision A A culture matrix, is characterized in that, include in described method accurate calculating, accurate weighing, first few after how mix, seal, prepared by independent mother liquor, six steps such as use of breaking seal.

2. method according to claim 1, is characterized in that, six described step concrete operations are as follows:

1. accurately calculate: 16 kinds of elements contained by AA (carbonaceous sources and mounting medium) minimal medium are expanded 1000 times, i.e. calcium chloride dihydrate 440g, potassium dihydrogen phosphate 170g, epsom salt 370g, potassium chloride 2940g, potassium iodide 0.83g, boric acid 6.2g, Sodium Molybdate Dihydrate 0.25g, white vitriol 8.6g, four water manganese sulphate 22.3g, iron edetate 36.5g, inositol 100g, thiamine hydrochloride 0.5g, glycine 75g, Glu 877g, L-Aspartic acid 266g, L-arginine 228g, 2 kinds of element CoCL2 6H2Os and cupric sulfate pentahydrate expand 10000 times, i.e. each 0.25g,

3. method according to claim 1, is characterized in that, the scale preparation method of this high precision culture matrix has following characteristics, i.e. without the error of calculation between the matrix of different batches; Because 16 kinds of reagent all expand 1000 times, 2 kinds of reagent expand 10000 times, and the weighing error between the matrix of different batches is minimum, and precision is high; In theory, due to the usage amount (calculating with each 1L) that solid matrix is 1000 times at every turn, namely the above-mentioned formula of 1000L can be prepared, liquid mother liquor only needs 100 μ L at every turn, and batch inside there will not be the human error caused because using the reagent of different manufacturers, different times; Solid matrix after expansion is that every 250g is sealed in plastic containers, can avoid because the impact of the extraneous factor such as weather, temperature, moisture causes occurring the common precipitation of liquid mother liquor, the phenomenon such as mouldy, and fluid matrix is only the mixing of two kinds of elements; After expanding 10000 times, precision is high; Above-mentioned plurality of advantages is all the follow-up test carried out for matrix with this medium, obtains result of the test accurately and lays the foundation.