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High-frequency roegneria kamoji immature embryo callus induction and regeneration cultivation method

A technology of callus induction and culture method, applied in the biological field, can solve problems such as inability to obtain successful in vitro culture, and achieve the effect of inhibiting endophyte contamination

Inactive Publication Date: 2014-09-24
NANJING AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

For the tissue culture of Gooseberry, endophytes are the source of pollution, which must be suppressed by disinfection and adding fungicides to the medium, otherwise the success of in vitro culture cannot be obtained.

Method used

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  • High-frequency roegneria kamoji immature embryo callus induction and regeneration cultivation method
  • High-frequency roegneria kamoji immature embryo callus induction and regeneration cultivation method
  • High-frequency roegneria kamoji immature embryo callus induction and regeneration cultivation method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] Example 1: Inhibition of endophytic fungi in the process of callus induction by immature embryos of Goose View Grass

[0025] The immature seeds 12-15 days after the flowering of Goose View Grass are peeled off from the young ears ( figure 1 ), soak in 70% alcohol for 30s, then soak in 0.1% mercuric chloride and shake for 15min, rinse with sterile water 3-5 times. Under sterile conditions, the immature embryos were stripped under a dissecting microscope and inoculated into induction medium MSD2 and MSD2 supplemented with 15mg / L Benlate. After 10 days of dark culture, count the number of embryos contaminated by endophytic fungi. In the MSD2 medium, the contamination rate was as high as 66.7%, but in the MSD2 medium supplemented with 15 mg / L Benlate, the contamination rate dropped to 3.3% (Table 1). Therefore, adding 15mg / L Benlate to the medium can effectively inhibit the growth of endophytic fungi.

[0026] The MSD2 medium is prepared by directly adding 2 mg / L of the ...

Embodiment 2

[0029] Example 2: Comparison of differences in callus induction rate of immature embryos of different sizes

[0030] Peel off the immature seeds 12-15 days after flowering of Goose View Grass from the young ears, soak in 70% alcohol for 30 seconds, then soak in 0.1% mercury chloride and shake for 15 minutes, and rinse with sterile water 3-5 times. Under aseptic conditions, the immature embryos were peeled off under a dissecting microscope, and were divided into three types according to the length of the longitudinal axis of the immature embryos ( figure 2 ): L stands for immature embryos with a length of about 300um, M stands for immature embryos with a length of about 200um, and S stands for immature embryos with a length of about 100um, which are respectively inoculated into MSD2 with 15mg / L Benlate added to the induction medium, cultured in the dark After 28 days, count and compare the differences in the callus induction rate of immature embryos of different sizes, and the...

Embodiment 3

[0033] Example 3: Effects of adding different hormone combinations on induction of immature embryo callus in induction medium

[0034] The 100-200um immature embryos of A. argentina in the above-mentioned Example 2 were selected for callus induction, and the effects of adding different hormone combinations (Table 3) in the MS induction medium on the callus induction of immature embryos were compared. Table 4 shows the healing rate and direct germination rate of immature embryos in MS medium supplemented with different hormone combinations. It can be seen from these two tables that the recovery rate of immature embryos in the culture without hormone 6-BA is higher than that in the medium with hormone 6-BA, while the overall growth rate of immature embryos in the medium with the hormone dicamba is The healing rate is better than that of the medium supplemented with hormone 2,4-D. Looking at the healing rate of immature embryos in all media, it is found that the healing rate of ...

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Abstract

The invention belongs to the field of biotechnology and relates to a high-frequency roegneria kamoji immature embryo callus induction and regeneration cultivation method. The method mainly comprises the following steps of: by taking immature seeds of roegneria kamoji as materials, sterilizing and stripping young embryo, arranging the young embryo on an induction medium, and cultivating to obtain the callus, performing high-frequency induction on buds in the bud initiation culture medium, and obtaining a regenerated plant in strong plantlet and rootage culture medium. In the whole culture stage, benomyl is added, so that the growth of endophytic fungi can be effectively inhibited. The culture of the callus is induced, 2,4-D, Dicamba and 6-BA according to a certain ratio is added, so that the induction rate of the callus can be effectively improved to be over 75 percent. The concentration ratio of the plant growth regulator is changed from the differentiation culture to regeneration culture, the bud induction rate is over 90 percent, and the plant regeneration rate is over 85 percent.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to a high-frequency culture method for inducing and regenerating callus of immature embryos of A. Background technique [0002] In order to meet the food demand of the ever-increasing population, people are urged to plant large-scale wheat varieties with a relatively single genetic background, which narrows the genetic basis of cultivated wheat, reduces genetic variation, and weakens resistance to diseases and insect pests and environmental adaptability. In order to solve this outstanding problem, breeders began to turn their attention to close relatives of wheat with rich genetic resources, hoping that through distant hybridization, beneficial genes of different species of plants could be introduced into cultivars, and the economical value of cultivars could be improved and improved. traits, especially improving the ability to resist biotic and abiotic stress, and cultivating crop variet...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A01H4/00
Inventor 邢莉萍钱晨王苏玲曹爱忠陈佩度
Owner NANJING AGRICULTURAL UNIVERSITY
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