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Young embryo transformation method of corn selfing line SL1303

An inbred line and corn technology, applied in the field of crop genetics and breeding, can solve problems such as imperfect transformation conditions

Active Publication Date: 2018-12-21
YANGZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to structural characteristics of callus and imperfect transformation conditions, there is no report on Agrobacterium-mediated genetic transformation using callus.

Method used

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  • Young embryo transformation method of corn selfing line SL1303
  • Young embryo transformation method of corn selfing line SL1303
  • Young embryo transformation method of corn selfing line SL1303

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] Embodiment 1: experimental material

[0046] 1 plant material

[0047] The young embryos harvested in the same year of the maize inbred line SL1303 came from the Maize Breeding Group of the Agricultural College of Yangzhou University, and were planted and preserved in our laboratory (Gao et al, 2016).

[0048] 2 experimental reagents

[0049] MS and N6 medium powder, 2,4-D (2,4-dichlorophenoxyacetic acid) and IBA (indole butyric acid) were purchased from Shanghai Sangon Bioengineering Technology Service Company.

[0050] 3 media

[0051] The basic medium is MS and N6 medium, both of which contain five components including trace elements, macroelements, iron salts, calcium salts and organic matter, but the content of each component is different. 2,4-D (2,4-dichlorophenoxyacetic acid) added to the basic medium induced embryogenic callus; CA (casamino acid) was used as an organic nutrient to promote callus growth and differentiation; KT (6- furfuryl aminopurine) and NA...

Embodiment 2

[0054] Example 2: Effects of Different 2,4-D Concentrations on Maize Callus

[0055]Three 2,4-D concentrations of 2, 4, and 6 mg / L were designed to cultivate callus, and 30 corn embryos of SL1303 were placed in each concentration, and three replicates were set for each concentration. After the callus was induced for 10 days, the callus length was counted. Bud rate, subculture the callus, observe the change of the callus state during the subculture process, and select the 2,4-D that can most inhibit the production of endogenous buds and is most likely to produce embryogenic callus from the three concentrations concentration.

[0056] It can be seen from Table 2 that within a certain concentration range, the increase of 2,4-D concentration can promote the inhibition of endogenous sprouts. When the concentration of medium 2,4-D was 6.0mg / L, the inhibitory effect on endogenous sprouts was the best, and the sprouting rate was only 28.89%. Different concentrations of 2,4-D induced...

Embodiment 3

[0065] Embodiment 3: rooting medium

[0066] The rooting medium was designed, a single aseptic seedling was cut from the SL1303 callus, and transplanted into different rooting mediums, one without IBA and one with 0.4 mg / L IBA. research shows( Figure 4 ), after the seedlings were transferred to the rooting medium that added 0.4mg / L IBA, they could produce 0.5-1.0cm thick young roots and grow rapidly in 3-4 days, and they could be hardened and transplanted in about 10d, which is better than Plus IBA medium is more suitable for rooting medium.

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Abstract

The invention belongs to the field of genetic breeding of crops, and particularly relates to a young embryo transformation method of a corn selfing line SL1303. The young embryo transformation methodcomprises the following steps of (1) inducing of callus: stripping the young embryos on a sterile operation table, using a pair of tweezers to fix corn seeds, picking out the embryo of the corn, slicing into two halves along the longitudinal axis of the embryo, and putting into an inducing culture medium for inducing culture; (2) subculture: cutting the endogenous bud out of the induced callus, inoculating into a subculture medium, culturing for seven days at the temperature of 25 DEG C under the dark environment, passaging for two generations, and calculating the grown embryonic callus; (3) differentiation culture: transferring the grown embryonic callus into a regeneration culture medium, radiating by light at the temperature of 25 DEG C, and culturing until the bud grows; (4) rooting culture: after the complete regenerated seedling is grown, transferring into a rooting culture medium, performing rooting and seedling exercising, and performing seedling exercising and transplanting after the root grows to about 4cm. The high-efficiency young embryo transformation method of the corn selfing line SL1303 is constructed, so as to lay a solid foundation for using the system to completethe transgenosis of the corn selfing line SL1303 in future.

Description

technical field [0001] The invention belongs to the field of crop genetics and breeding, and in particular relates to a method for transforming immature embryos of corn inbred line SL1303. At present, the Agrobacterium-mediated method is the most commonly used method for crop transgenic research, and the key to successfully obtaining transgenic plants is the establishment of an efficient and stable genetic transformation and regeneration system. Immature embryos are the earliest and most successful explants in maize tissue culture. Genetic transformation of immature embryos as recipient materials has good somatic cell reproduction and regeneration effects. The establishment of a transformation system using immature embryos as recipient materials is of great significance. important meaning. Background technique [0002] 1 Introduction [0003] Maize (Zea mays L.) is the most widely planted cereal crop with the highest yield in the world. It has multiple uses such as grain, ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01H1/02A01H4/00
CPCA01H1/02A01H4/001A01H4/008
Inventor 高勇陈建民陶海霞仲雪张梦娇
Owner YANGZHOU UNIV
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