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Efficient transformation method for transforming agrobacterium into setaria viridis seeds under assistance of carborundum

A technology of Agrobacterium transformation and carborundum, applied in the biological field, can solve the problems of reduced transformation efficiency of embryogenic callus, reduced flexibility and simple practicability, and long induction time of embryogenic callus, so as to shorten the transformation period, The effect of short cycle and easy operation

Inactive Publication Date: 2017-05-31
INST OF TROPICAL BIOSCI & BIOTECH CHINESE ACADEMY OF TROPICAL AGRI SCI
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  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, the transgenic technology of Setaria mainly adopts the method of inducing embryogenic callus from the meristems of the young bud nodes after mature seeds germinate to carry out Agrobacterium transformation. The biggest problem with this method is that the induction rate of embryogenic callus is low. Different from the condition of seeds, the ratio of the number of induced embryogenic callus to the number of seeds is generally less than 10%. Secondly, the induction time of embryogenic callus is longer, and a small amount of embryogenic callus in the first stage is obtained from the inoculation to the medium. It takes about one month for the wounded tissue, and then it needs to be subcultured twice (once every half month) before there is enough embryogenic callus for Agrobacterium transformation, and the whole transformation cycle needs about 4 months; meanwhile, this Embryogenic callus is not resistant to subculture and cannot be subcultured for long-term preservation for transformation. The transformation efficiency of embryogenic callus subcultured more than 3 times is significantly reduced, and it is not suitable for continued transformation; in addition, embryogenic callus The selection requires some experience
These all reduce the flexibility and simple practicality of this conversion technology

Method used

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  • Efficient transformation method for transforming agrobacterium into setaria viridis seeds under assistance of carborundum
  • Efficient transformation method for transforming agrobacterium into setaria viridis seeds under assistance of carborundum
  • Efficient transformation method for transforming agrobacterium into setaria viridis seeds under assistance of carborundum

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Experimental program
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Embodiment

[0026] 1. Treatment and activation of explants

[0027] Select dried and mature seeds of Setaria lines A10 and ME34 after dormancy treatment, after removing the seed coat, put about 1 gram of each seed into two 14ml centrifuge tubes, sterilize and disinfect with 1% effective concentration of sodium hypochlorite solution, and wash with sterile water Finally, add about half the volume of bud induction liquid medium in the centrifuge tube and shake it for 5 days. The culture conditions: light culture, and the culture temperature is about 24 ° C. The bud induction liquid medium is based on the modified MS medium (trace element content is doubled), the pH is 5.8, and 2mg / L2,4-D+0.5mg / L KT is added.

[0028] 2. Emery wound treatment and Agrobacterium infiltration

[0029] According to the seed volume ratio (1:1), corundum (half the weight of 120 mesh and half of 320 mesh) was weighed, subjected to high-temperature sterilization, and the bud induction liquid medium in the centrifuge...

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Abstract

The invention belongs to the field of biotechnology and relates to an efficient transformation method for transforming agrobacterium into setaria viridis seeds under assistance of carborundum. The efficient transformation method for transforming the agrobacterium into the setaria viridis seeds under assistance of the carborundum comprises the processes of explant treatment and activation, carborundum wound treatment and agrobacterium impregnation, co-culture, bacteriostasis and screening culture, resistant callus seedling culture, rooting culture and so on. The efficient transformation method for transforming the agrobacterium into the setaria viridis seeds under assistance of the carborundum, provided by the invention, has the advantages that the steps of the method are simple; the method is easy to operate; as the mature setaria viridis seeds are used as explants and wound are formed on the seeds under the assistance of the carborundum to carry out agrobacterium transformation after the seeds germinate and are activated, the transformation efficiency of the setaria viridis is greatly enhanced, the transformation period is reduced, the transformation procedure is simplified and the transformation strain range is enlarged; and a flexible and efficient transgenic technology system with short period is established for the setaria viridis.

Description

technical field [0001] The invention belongs to the field of biological technology, and relates to a high-efficiency transformation method of Agrobacterium setariae, in particular to a high-efficiency transformation method for Agrobacterium carborundum to transform Setaria seeds. Background technique [0002] C4 photosynthesis drives the productivity of several major food crops and bioenergy grasses, including maize (Zeamays), sugarcane (Saccharum officinarum), sorghum (Sorghum bicolor), Miscanthus giganteus, and switchgrass (Panicum virgatum), among others. Setaria viridis is a true diploid with many characteristics suitable for genetic analysis, including a relatively small genome (510Mb), short stature, short life cycle and large seed production, etc. An excellent model for C4 photosynthesis, but also for studying abiotic stress tolerance and bioenergy feedstocks. Morphologically similar to most grasses, including major bioenergy crops switchgrass (Panicum virgatum) and ...

Claims

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Application Information

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IPC IPC(8): C12N15/82A01H5/00
Inventor 赵辉郭静远郭安平
Owner INST OF TROPICAL BIOSCI & BIOTECH CHINESE ACADEMY OF TROPICAL AGRI SCI
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