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A kind of method of wheat mature embryo culture and regeneration

A mature embryo culture and mature embryo technology is applied in the field of callus induction and regeneration of wheat mature embryos, which can solve the problems of low average level of differentiation rate and achieve the effects of increasing concentration, reducing cost and low price.

Inactive Publication Date: 2011-12-21
JIANGSU ACADEMY OF AGRICULTURAL SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] The purpose of the present invention is to provide a new method for culturing and regenerating wheat mature embryos, aiming at the fact that the average level of callus differentiation rate of mature wheat embryos is relatively low at present.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] 1) Induction medium

[0019] The induction medium contains the macroelements, trace elements, iron salts of MS basic medium, and vitamins of B5 basic medium, adding maltose 40g / L, magnesium chloride 0.75g / L, hydrolyzed casein 500mg / L, and glutamine 300mg / L, MES 1.95g / L solid medium, pH=5.8.

[0020] After preparation, it was sterilized by high pressure damp heat at 121°C for 20 minutes.

[0021] 2) Regeneration medium (referred to as KT 5.0NAA 0.1 regeneration medium)

[0022] The regeneration medium is a solid medium containing MS basic medium, maltose 40g / L, MES 1.95g / L, KT 5mg / L, NAA 0.1mg / L, pH=5.8.

[0023] After preparation, it was sterilized by high pressure damp heat at 121°C for 20 minutes.

[0024] 3) Regeneration medium for control (referred to as KT 5.0 regeneration medium)

[0025] The regeneration medium used for the control is the basic medium containing MS, 40 g / L maltose, 1.95 g / L MES, 5 mg / L KT, pH=5.8.

[0026] After preparation, it was steriliz...

Embodiment 2

[0028] Wheat variety "Huapei No. 1".

[0029] Pick mature and plump "Huapei No. 1" wheat seeds, soak them in 70% alcohol for 5 minutes, and use 0.1% HgCl 2 Sterilize for 20 minutes, rinse with sterile water for 4 times, soak in sterile water for 19 hours, and obtain mature embryos of the seeds.

[0030] After the mature embryo of wheat seeds was removed from the radicle, it was longitudinally cut and inserted into the above-prepared induction medium (provided in Example 1, the same below), cultured in the dark at 23°C, subcultured once every 10 days, and co-induced for 20 days to obtain Wheat callus.

[0031] The wheat callus was transferred to KT 5.0 NAA 0.1 regeneration medium and KT 5.0 regeneration medium (both provided by Example 1, the same below), respectively, to induce the differentiation of regenerated shoots. The culture period is 30 days. During the culture period, the ambient temperature is 23-25° C., light and darkness are alternated every 12 hours, and the lig...

Embodiment 3

[0035] Wheat variety "Ningmai 13"

[0036] Pick mature and plump "Ningmai 13" wheat seeds, soak them in 70% alcohol for 5 minutes, and use 0.1% HgCl2 Sterilize for 20 minutes, rinse with sterile water for 5 times, and soak in sterile water for 24 hours to obtain mature embryos of seeds.

[0037] The mature embryos of wheat seeds were cut longitudinally after removing the radicle radicle, and inserted into the induction medium prepared above, cultured in the dark at 25°C, subcultured once every 14 days, and co-induced for 28 days to obtain wheat callus.

[0038] The wheat callus was transferred to KT 5.0 NAA 0.1 regeneration medium and KT 5.0 regeneration medium to induce the differentiation of regenerated shoots. The culture period is 28 days. During the culture period, the ambient temperature is 23-25° C., light and darkness are alternately induced every 12 hours, and the light intensity is 2000 lux. The mature wheat embryo callus differentiates into regenerated plant shoots....

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Abstract

The invention relates to a mature wheat embryo culture and regeneration method. Regeneration culture mediums are based on MS basic culture mediums. In the regeneration culture mediums, the content of maltose is 40g / L, the content of MES is 1.95g / L, the content of KT is 5mg / L, the content of NAA is 0.1mg / L and the pH is 5.8. The specific method comprises the steps of selecting mature and full wheat seeds, soaking the seeds in 70% alcohol solution for 5min, sterilizing the seeds in 0.1% HgCl2 solution for 20min, washing the seeds for at least four times by using sterile water and soaking the seeds in the sterile water for 19-24h; removing the buds and the roots of the mature embryos of the seeds, longitudinally cutting the mature embryos, putting the cut mature embryos into induction culture mediums to culture in the dark at 23-25 DEG C, conducting subculture once for 10-14 days and conducting induction culture for totally 20-28 days to obtain mature wheat embryo calluses; and then transferring the mature wheat embryo calluses into the prepared regeneration culture mediums to culture in the light at 23-25 DEG C for 25-30 days and inducing the mature wheat embryo calluses to be differentiated to form regenerated seedlings.

Description

technical field [0001] The invention relates to a method suitable for induction and regeneration of mature wheat embryo callus. Background technique [0002] Wheat tissue culture provides a necessary basis for wheat clonal variation, embryo rescue, and transformation of exogenous genes, and high-frequency regeneration is the key to wheat tissue culture. [0003] At present, the successful explants of wheat tissue culture include roots, young ears, immature embryos, anthers, and mature embryos. Among them, the regeneration efficiency of immature embryo culture is the highest, while the regeneration of mature embryo culture is the most difficult. However, the selection of immature embryos is greatly limited by seasons, time, and different physiological states of individual plants. Under natural conditions, it cannot be continuously supplied throughout the year. The use of greenhouse cultivation techniques for continuous planting throughout the year requires strict conditions ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01H4/00
Inventor 蒋苏马鸿翔张旭余桂红孙晓波魏芳
Owner JIANGSU ACADEMY OF AGRICULTURAL SCIENCES
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