Method for improving agribacterium-mediated transformation survival rate of wheat

A technology of Agrobacterium and wheat, applied in biochemical equipment and methods, botany equipment and methods, applications, etc., can solve problems such as affecting transformation efficiency and death of wheat transformed seedlings, and achieve improved survival efficiency, large application value and market Prospect of promotion, low price effect

Active Publication Date: 2014-09-03
BEIJING WEIMING KAITUO CROP DESIGN CENT COMPANYLIMITED
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] However, when using longitudinally cut wheat seedling leaf bases for Agrobacterium transformation, the massive growth of mold during the co-cultivation process led to the death of most of the transformed wheat seedlings. This phenomenon is very common, which seriously affects the transformation efficiency.

Method used

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  • Method for improving agribacterium-mediated transformation survival rate of wheat
  • Method for improving agribacterium-mediated transformation survival rate of wheat

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] 1. Experimental materials

[0023] Plasmid: pV1 vector containing paraquat resistance gene and antibiotic selection marker gene, with paraquat resistance. The PCR amplified gene was ligated to pcambia1300 (purchased from Invitrogen) vector to obtain pV1 vector.

[0024] Agrobacterium strain: AGL0

[0025] Wheat variety for transformation: Jingmai 9158

[0026] 2. Cultivation of Agrobacterium

[0027] Pick the Agrobacterium containing the target gene cryopreserved in glycerol, draw a line on the YEB plate medium, and culture for 2 days in a dark incubator at 28°C. Pick a single colony and inoculate it in 40ml of resistant YEB liquid medium, shake it overnight at 28°C in a dark shaker at 200rpm, to OD 600 ≈1. Pipette 400μl of the above bacterial liquid and spread it on the YEB solid medium with a diameter of 15cm. After incubating in a dark incubator at 28°C for 2 days, do not continue to store it in a dark incubator at 28°C or transfer to a refrigerator at 4°C.

[0028] 3. Prepa...

Embodiment 2

[0066] 1. Experimental materials

[0067] Plasmid: pV1 vector containing paraquat resistance gene and antibiotic selection marker gene. The PCR amplified gene was ligated to pcambia1300 (purchased from Invitrogen) vector to obtain pV1 vector.

[0068] Agrobacterium strain: AGL0

[0069] Wheat variety for transformation: Jingmai 9158

[0070] 2. Cultivation of Agrobacterium

[0071] Pick the Agrobacterium containing the target gene cryopreserved in glycerol, draw a line on the YEB plate medium, and culture for 2 days in a dark incubator at 28°C. Pick a single colony and inoculate it in 40ml of resistant YEB liquid medium, shake it overnight at 28°C in a dark shaker at 200rpm, to OD 600 ≈1. Pipette 400μl of the above bacterial liquid and spread it on the YEB solid medium with a diameter of 15cm. After incubating in a dark incubator at 28°C for 2 days, do not continue to store it in a dark incubator at 28°C or transfer to a refrigerator at 4°C.

[0072] 3. Preparation of conversion solut...

Embodiment 3

[0111] 1. Experimental materials

[0112] Plasmid: pV1 vector containing paraquat resistance gene and antibiotic selection marker gene. The PCR amplified gene was ligated to pcambia1300 (purchased from Invitrogen) vector to obtain pV1 vector.

[0113] Agrobacterium strain: AGL0

[0114] Wheat variety for transformation: Jingmai 9158

[0115] 2. Cultivation of Agrobacterium

[0116] Pick the Agrobacterium containing the target gene cryopreserved in glycerol, draw a line on the YEB plate medium, and culture for 2 days in a dark incubator at 28°C. Pick a single colony and inoculate it in 40ml of resistant YEB liquid medium, shake it overnight at 28°C in a dark shaker at 200rpm, to OD 600 ≈1. Pipette 400μl of the above bacterial liquid and spread it on the YEB solid medium with a diameter of 15cm. After incubating in a dark incubator at 28°C for 2 days, do not continue to store it in a dark incubator at 28°C or transfer to a refrigerator at 4°C.

[0117] 3. Preparation of conversion solut...

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Abstract

The invention discloses a new method for transforming wheat by longitudinally cutting the leaf base of seedlings. When using the longitudinally cutting seedling leaf base for Agrobacterium transformation, an antiseptic is added to the co-cultivation substrate, thereby restraining the growth of mold and improving the Survival of transformed plants.

Description

technical field [0001] The invention belongs to the field of wheat transgenes, and relates to a method for improving the survival efficiency of wheat by using a preservative to transform the base of longitudinally cut seedling leaves mediated by Agrobacterium, and further improving the transformation efficiency. Background technique [0002] Wheat is the second most important food crop in terms of total output and is widely planted all over the world. With the growth of the world population and the improvement of people's living standards, the demand for wheat is increasing, and the requirements for wheat quality are also getting higher and higher. Conventional breeding can no longer meet people's requirements for wheat varieties and quality. The development of transgenic technology has broken the racial restrictions of conventional breeding. Many crops such as corn and rapeseed have successfully bred high-quality and high-efficiency transgenic new varieties using transgenic...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/84A01H5/00
Inventor 冉冬夏勉张斌何峰郑辰
Owner BEIJING WEIMING KAITUO CROP DESIGN CENT COMPANYLIMITED
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