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Cultivation method of transgenic Lotus corniculatus tissue culture seedlings

A cultivation method and technology of japonicus japonicus, applied in the field of cultivation of transgenic japonicus japonicus tissue culture seedlings, achieving the effects of low cost, high transformation efficiency, and large amplification coefficient

Inactive Publication Date: 2018-09-14
SICHUAN AGRI UNIV
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Problems solved by technology

However, it is worth noting that so far, there have been few research reports on calmodulin family proteins in rhizobium-legume symbiotic nitrogen fixation, such as which calmodulin family proteins in lotus root are involved in rhizobium-legume symbiotic nitrogen fixation. Symbiotic nitrogen fixation, how the expression mechanism is rarely reported

Method used

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Experimental program
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Embodiment 1

[0036] A method for cultivating transgenic lotus root tissue culture seedlings, comprising the following steps:

[0037] (1) Seed pretreatment

[0038] Since the seed coat of japonicus japonicus is thick and impermeable, the seeds can be gently rubbed with sandpaper to polish the seed coat lightly, which is conducive to the rapid germination of the seeds.

[0039] (2) Seed disinfection

[0040] Immerse the pretreated seeds in a mixed solution of 70% ethanol and 1% SDS by volume fraction, sterilize for 5min, remove ethanol, and then use 20% NaClO and 0.1% NaClO with mass concentration. The mixed solution of SDS is used to sterilize the seeds for 10 minutes. After the disinfection is completed, take out the seeds, rinse the sterilized seeds with sterile MilliQ water for 15-20 times in a sterile ultra-clean workbench, and finally soak the washed seeds in in sterile MilliQ water overnight at room temperature.

[0041] (3) Germination of seeds

[0042] Spread 0.5cm thick steril...

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Abstract

The invention discloses a cultivation method of transgenic Lotus corniculatus tissue culture seedlings. The method includes the steps of firstly, preprocessing and sterilizing seeds; secondly, allowing the seeds to germinate; thirdly, performing agrobacterium-mediated transformation and cultivation; fourthly, performing co-cultivation; fifthly, performing rebirth; sixthly, performing selective cultivation; seventhly, performing bud induction and extension; eighthly, performing root induction and extension. The method has the advantages that Lotus corniculatus calmodulin genes are connected toplasmids, then Lotus corniculatus is transformed through agrobacterium mediation, and transgenic tissue culture experiments are used to obtain overexpression Lotus corniculatus plants; the method is low in cost, simple to operate and high in survival rate and transformation rate.

Description

technical field [0001] The invention belongs to the technical field of cultivation of transgenic plants, and in particular relates to a method for cultivating transgenic japonicus japonicus tissue culture seedlings. Background technique [0002] Biological nitrogen fixation plays a very important role in agricultural production. It can provide nitrogen for food crops, increase yield, reduce fertilizer consumption and production costs, reduce environmental pollution, prevent land desertification, establish ecological balance and promote sustainable agricultural development. Legumes can form root nodules in symbiosis with rhizobia in the soil, and the root nodules can fix nitrogen in the air and provide it for plant growth and development. Root nodule nitrogen fixation must meet two conditions, one is that rhizobia enter root epidermal cells through root hairs to form invasion threads (infection threads, IT), that is, the genetic pathway of epidermal cells; nidulation primodi...

Claims

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Application Information

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IPC IPC(8): A01H4/00
CPCA01H4/001A01H4/008
Inventor 唐学博秦宗志廖进秋
Owner SICHUAN AGRI UNIV
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