Oncidium protoplast dissociation and culture method

A technology of protoplast and culture method, applied in the field of ornamental plant cell engineering, can solve the problem that oncidium protoplast has not been reported, and achieve the effect of good free effect

Pending Publication Date: 2020-06-16
陈俊敏
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the research on the isolation and cultivation of orchid protoplasts mainly focuses on Cymbidium, Phalaenopsis, and Dendrobium, but the research on Oncidium protoplasts has not been reported yet.

Method used

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  • Oncidium protoplast dissociation and culture method
  • Oncidium protoplast dissociation and culture method
  • Oncidium protoplast dissociation and culture method

Examples

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Embodiment 1

[0027] 1 Induction and subculture of callus

[0028] Induction of callus: cut oncidium protocorms into small pieces of 1-2 mm, inoculate them on callus induction medium, and culture them in the dark at 25°C for about 30 days; The formula is: VW basic medium + Huabao No. 1 3g / L + 6-BA1.0mg / L + NAA0.1mg / L + ferulic acid 10mg / L + sucrose 40g / L + agar 6g / L, adjust the pH of the medium to 5.8 .

[0029] Subculture of callus: transfer the induced callus to the subculture medium for cultivation, the temperature is 25°C, the light intensity is 800lx, select subculture every 15~20d, subculture 4~5 times Obtain a callus with vigorous growth and loose structure for the dissociation of protoplasts; the formula of the subculture medium is: VW basic medium + Huabao No. 1 3g / L+NAA3.0mg / L+IAA 2.0mg / L+ 6-BA0.2mg / L + sucrose 30g / L + agar 6g / L; adjust the pH of the medium to 5.8.

[0030] 2 Isolation of protoplasts

[0031] Isolation of protoplasts: cold-treat the callus obtained from subcul...

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Abstract

The invention provides an oncidium protoplast dissociation and culture method which comprises the following steps: taking oncidium protocorm-like bodies as starting materials, carrying out callus induction culture and 4-5 times of subculture to obtain vigorously growing calluses with loose structures for dissociation of protoplasts; carrying out cold treatment on calluses at 4 DEG C for 1 d, putting the calluses into a mixed enzyme solution, carrying out shaking table oscillation enzymolysis for 8-10 h, and purifying protoplasts by using an interface method to obtain a large amount of oncidiumprotoplasts with high activity; continuously using the culture medium for solid-liquid double-layer culture, so that the obtained oncidium protoplast cells can normally grow and continuously split to form small cell clusters, the maximum splitting frequency reaches 10.16%, and the maximum plate planting rate reaches 4.03%. The invention provides a scientific basis for cell fusion, somatic cell hybridization, genetic transformation and germplasm innovation by utilizing oncidium protoplast.

Description

technical field [0001] The invention belongs to the field of ornamental plant cell engineering, and in particular relates to a method for isolating and cultivating oncidium protoplasts. Background technique [0002] Oncidium ( Oncidium hybridum ), also known as auspicious orchid, dancing orchid, dancing girl orchid, King Phalaenopsis, tumor petal orchid, etc., is the general term for hybrids between the genus Oncidium and its close relatives, mainly distributed in the United States, Mexico, Paraguay, Peru, Brazil tropical and subtropical regions of Central and South America. Oncidium is a compound-stem aerial orchid with large morphological changes. It can be divided into species with pseudobulbs and species without pseudobulbs according to whether it has pseudobulbs. According to the shape of leaves, it can be divided into thin-leafed species, thick-leaved species There are three types of leaf species and sword leaf species. The flowers of oncidium vary greatly in shape ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/04
CPCC12N5/04C12N5/0025
Inventor 陈俊敏
Owner 陈俊敏
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