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LarixLeptolepis somatic embryo regeneration method

A technology of somatic embryos and larch, applied in the fields of plant regeneration, horticultural methods, botanical equipment and methods, etc. Induction efficiency, great practical application value, and the effect of reducing deformed embryos

Inactive Publication Date: 2020-06-16
BEIJING FORESTRY UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The object of the present invention is to provide a method for regeneration of larch somatic embryos, which solves the problems of low incidence of somatic embryos, long formation time of somatic embryos, high deformity rate, asynchronous occurrence, and low germination rate

Method used

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  • LarixLeptolepis somatic embryo regeneration method
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  • LarixLeptolepis somatic embryo regeneration method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] Example 1. Collect zygotic embryos from immature seeds of larch and induce embryogenic callus

[0034] Collect the immature seeds of Larix olgensis in mid-to-late June, soak in water for 4-6 hours and clean them, sterilize with 75% alcohol for 3 minutes, and sterilize with 5% sodium hypochlorite for 15 minutes, and then repeatedly clean with sterile distilled water and peel off the seed coat , Cut the endosperm to the embryo or directly remove the immature embryo, and inoculate the embryogenic callus induction medium for embryogenic callus induction (23±1℃, dark culture). After induction for about 30 days, the embryogenic calli were picked out, separated into small pieces, and proliferated and subcultured in the subculture medium for a long time (23±1℃, dark culture, subculture once every 15 days).

Embodiment 2

[0035] Example 2. Medium configuration

[0036] The composition of the embryogenic callus induction medium is as follows, which is an additional ingredient 1 / 2SPE medium (Gupta&Durzan, 1987, hereinafter referred to as S medium), plus agar 3g / L, the balance is water, pH5.8:

[0037] Table 1. Embryogenic callus induction medium composition

[0038]

[0039]

[0040] The composition of the subculture medium for embryogenic callus is as follows, which is the additional component of S medium, plus 5g / L agar, the balance is water, pH 5.8:

[0041] Table 2. Substitution medium composition of embryogenic callus

[0042] ingredient Content (mg / L) ingredient Content (mg / L) 2,4-D0.15KH 2 PO 4

85 6-BA0.05CaCl 2 ·2H 2 O

327 KT0.05MgSO 4 ·7H 2 O

172 Glutamine500H 3 BO 3

3.1 Hydrolyzed Casein500ZnSO 4 ·7H 2 O

4.3 Glycine2MnSO 4 ·H 2 O

8.45 sucrose30000Na 2 MoO 4 ·2H 2 O

0.125 Inositol1000 KI 0.415 Vitamin B 1

0.5CuSO 4 ·5H 2 O

0.0125 Vitamin B 6

0.5CoCl 2

0.0125 KNO 3

2337FeSO ...

Embodiment 3

[0052] Example 3. Regeneration of larch somatic embryos

[0053] 1. Liquid to solid medium culture method

[0054] The method includes the following steps:

[0055] 1. Using the zygotic embryos of immature seeds of Larix olgensis collected from June 15 to June 25 as explants, the embryogenic callus induced by Larix olgensis was the initial material for this experiment.

[0056] 2. Under aseptic operation conditions in the ultra-static workbench, select the embryogenic callus that has grown well and is cultured on the new subculture medium for about 10 days, and use tweezers to tear it up and connect it to the liquid medium A in an Erlenmeyer flask. , Each 1g callus is inoculated in about 50ml culture medium, cultured in the dark for 4 days under suspension conditions at 100rpm, 23±1℃, and suspension.

[0057] 3. Under aseptic operating conditions in an ultra-static workbench, use a fine filter to filter out the culture in Medium A and transfer it to an Erlenmeyer flask with a medium vo...

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Abstract

The invention provides a larixLeptolepis somatic embryo regeneration method, which comprises the following steps of: collecting a larixLeptolepis immature seed zygotic embryo as an explant, and inducing a larixLeptolepis embryonic callus; culturing for 7-10 days on a new subculture medium, selecting the embryonic callus which favorably grows, inoculating the embryonic callus in a liquid culture medium in a conical flask, and carrying out dark culture in a table concentrator at 50-125 rpm and a temperature of 20-25 DEG C under a suspension condition for 4-7 days; then, transferring a culture into a semisolid culture medium, and carrying out the dark culture in the table concentrator at 50-125 rpm and the temperature of 20-25 DEG C under the suspension condition for 4-7 days; and finally, inculcating the culture to the surface of a solid culture medium, carrying out the dark culture for 23-30 days at the temperature of 20-25 DEG C, and forming a later-stage cotyledon embryo. According tothe method, a great quantity of high-quality somatic embryos can be generated by induction, abnormal embryos are reduced, somatic embryo formation induction time is short, in addition, a better synchronization effect can be achieved, and induction efficiency is high.

Description

Technical field [0001] The invention relates to the field of larch plant cultivation, in particular to a method for regeneration of larch somatic embryos. Background technique [0002] Larix Leptolepis is a fast-growing coniferous wood species in northern and subtropical and mid-alpine areas of my country. It has the characteristics of wide distribution, strong adaptability, few diseases and insect pests, and excellent material. Its early rapid growth is among the top conifers. It is an afforestation tree species that both state-owned forest farms and farmers attach great importance to, and the area of ​​artificial forestation is expanding. Therefore, carrying out research work on larch breeding is an important field for improving coniferous trees in my country. The conventional propagation method of larch is seed propagation or asexual (cutting) propagation. However, due to the long propagation cycle of larch seeds, low germination rate, low asexual cutting propagation coefficie...

Claims

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Application Information

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IPC IPC(8): A01H4/00
CPCA01H4/001A01H4/008
Inventor 盖颖刘威锋吴晓雪蒋湘宁
Owner BEIJING FORESTRY UNIVERSITY
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