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A kind of genetic transformation method using mint internode stem segments as explants

A genetic transformation system and explant technology, applied in the directions of botanical equipment and methods, biochemical equipment and methods, plant genetic improvement, etc. effect of growth

Active Publication Date: 2021-11-30
INST OF BOTANY JIANGSU PROVINCE & CHINESE ACADEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the rapid propagation system for domestic mint varieties mainly uses noded stems as regeneration materials. Xie Wenshen et al. (2016) reported a tissue rapid propagation method for virus-free seedlings of wild mint. Nodal stem segment, Wu Tao et al. (1999) also reported the method of rapid propagation with a single segmental stem segment with leaves, but these methods are only suitable for large-scale multiplication of mint varieties and are not suitable for genetic transformation methods

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] Carry out the genetic transformation using the internode stem section of peppermint without nodes as explants in the following steps:

[0023] (1) Activation culture of bacteria:

[0024] Streak the Agrobacterium tumefaciens LBA4404 on the YEB solid medium, culture it in the dark at 28°C for 36 hours, pick a single colony and insert it into the YEB liquid medium, and shake it at 28°C for 2 days , the measured bacterial solution OD600 = 0.8, used to impregnate the internode stems of mint without nodes.

[0025] (2) Obtaining sterile vaccines:

[0026] Put the mint stem tip on the ultra-clean workbench, first disinfect it with 75% alcohol for 30 seconds, then sterilize it with 0.1% mercury liter for 10 minutes, rinse it with sterile water for 3 times, and peel off the most sterilized stem tip Inoculate MS+6-BA 2 mg·L after the outer 3 leaves -1 +NAA0.2 mg·L -1 + sucrose 30g·L -1 + agar 3.8 g·L -1 ;

[0027] (3) Pre-cultivation of mint internode stems without nodes:...

Embodiment 2

[0032] The process of this embodiment is basically the same as Embodiment 1, the difference is:

[0033] In step (1), the concentration of the Agrobacterium tumefaciens EHA105 bacteria liquid concentration OD600=0.8 used to infiltrate the internode stem explants of peppermint is used to infect the internode stem explants of mint. The regeneration rate of resistant shoots was 10.23% after 40 days of culture in the screening medium in step (5).

Embodiment 3

[0035] The process of this embodiment is basically the same as Embodiment 1, the difference is:

[0036] In the step (1), the concentration of Agrobacterium tumefaciens LBA4404 bacterium liquid concentration OD600=0.6 that infiltrates the internode stem explants of mint is impregnated and infects the internode stem explants of mint. The regeneration rate of resistant shoots was 10.44% after 40 days of culture in the selection medium in step (5).

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Abstract

The invention discloses a genetic transformation method using mint internode stem segments as explants. Precultivation of interstem segments, step 4, genetic transformation of mint internodes; step 5, rapid proliferation of resistant buds. By adopting the method, the genetic transformation of the mint stem segment is successfully realized, and stable genetic and fast-growing transgenic positive plants are obtained.

Description

[0001] The invention specifically relates to a field of peppermint genetic transformation technology, and particularly discloses a method for genetic transformation of peppermint internode stem segments into explants. Background technique [0002] Mint( Mentha haplocalyx Briq.) is rich in volatile oil in stems and leaves, and is widely used in food, medicine, cosmetics, spices, tobacco and other industries. The whole herb of mint is used as medicine to treat wind-heat cold, the beginning of wind and temperature, headache, red eyes, sore throat, sore throat, sore tongue, toothache, urticaria, rubella, etc. Mint is one of the important spices and medicinal plants in China. It is widely cultivated in Jiangsu, Anhui, Jiangxi and other places, and has become an important agricultural special economic crop in the local area. However, the phenomenon of single species and species degeneration in mint production has made mint breeding work more and more attractive. The use of transg...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/84
CPCC12N15/8205
Inventor 于盱梁呈元房海灵亓希武李维林尚庆文巢建国
Owner INST OF BOTANY JIANGSU PROVINCE & CHINESE ACADEMY OF SCI
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