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Tetraploid transgenic locust and quick cultivation

A technology of tetraploid Robinia pseudoacacia and tissue culture rapid propagation, which is applied in the field of plant genetic engineering to achieve high transformation efficiency

Inactive Publication Date: 2005-02-23
SHANDONG FOREST SCI RES INST
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, so far, there has not been any meaningful target gene introduced into Robinia pseudoacacia

Method used

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  • Tetraploid transgenic locust and quick cultivation
  • Tetraploid transgenic locust and quick cultivation
  • Tetraploid transgenic locust and quick cultivation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0049] The plant material is an excellent variety of feed-type tetraploid Robinia pseudoacacia introduced from Korea. The explants were collected from the axillary shoots of the excellent plants in the field, rinsed with tap water, sterilized with 70% alcohol, rinsed with sterile water for 3-5 times, then disinfected with mercuric chloride solution, rinsed with sterile water for 5 times, After being inoculated in the starting medium, the axillary buds of the stem segments will start to germinate after 12 days, and shoots will emerge after 20 days, which are used as callus induction materials.

[0050] Among them, the starting medium composition is MS+6-BA 0.3mg·L -1 +NAA 1.5mg·L -1 ; Callus induction medium is: MS+KT 1.5mg L -1 +NAA 1.5mg·L -1 ; The differentiation medium is: MS+IBA 1.0mg·L -1 +6-BA 3.5mg·L -1 . The composition of MS medium is: KNO 3 1900mg·L -1 , NH 4 NO 3 1650mg·L -1 , MgSO 4 370mg·L -1 , KH 2 PO 4 170mg·L -1 , CaCl 2 2H 2 O 440mg·L -1...

Embodiment 2

[0060] Tetraploid Robinia pseudoacacia explants (new shoots and stems), inoculated in the starting medium MS+6-BA 1.0mg L after disinfection -1 +NAA 1.0mg·L -1After 12 days, the axillary buds of the stem segment began to germinate, and after 15 days, the shoots were inoculated into the callus induction medium MS+KT 1.5mg·L -1 +NAA 2.0mg·L -1 , the obtained callus was then transferred to the differentiation medium MS+IBA 1.5mg·L -1 +6-BA 2.5mg·L -1 middle. The sucrose concentration of each medium is 25g·L -1 , agar 7.0g·L -1 , pH 5.8.

[0061] A single colony of Agrobacterium carrying the BADH gene was selected and inoculated in YEB liquid medium containing Kan, cultured overnight (25°C, 180rin / m), transferred to YEB medium without antibiotics, and the bacterial solution was centrifuged, then Add appropriate amount of sterile MS liquid medium to dilute to OD 600 0.5. Cut the callus into thin slices, and infect the Agrobacterium bacteria solution for 10 minutes after t...

Embodiment 3

[0064] Tetraploid Robinia pseudoacacia explants (new shoots and stems), inoculated in the starting medium MS+6-BA 1.5mg L after disinfection -1 +NAA 0.5mg·L -1 After 15 days, the axillary buds of the stem segment began to germinate, and the shoots were sent out after 18 days, and the shoots were inoculated into the callus induction medium MS+KT 2.5mg·L -1 +NAA 1.5mg·L -1 The callus was obtained from the medium, and then transferred to the differentiation medium MS+IBA 0.5mg·L -1 +6-BA 3.5mg·L -1 middle,. The sucrose concentration of each medium is 35g L -1 , agar 8.0g·L -1 , pH 5.8.

[0065] A single colony of Agrobacterium carrying the BADH gene was selected and inoculated in YEB liquid medium containing Kan, cultured overnight (30°C, 190rin / m), then transferred to YEB medium without antibiotics for culture, and the bacterial solution was centrifuged, then Add appropriate amount of sterile MS liquid medium to dilute to OD 600 0.6. Cut the callus into thin slices, an...

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Abstract

A transgenic and tissue culture method for quickly reproducing the tetraploid locust tree includes such steps as sampling the new stem of tetraploid locust tree, disinfecting, start culturing, inducing culture to obtain calli, transgenic treating by Agrobacterium tumefaciens LBA 4404 containing binary carrier pBin 438, differential culturing, selection culturing to obtain resistant plant Kan, molecular testing to obtain transgenic plant, rooting culture, naturalizing and planting in field.

Description

(1) Technical field [0001] The invention relates to a breeding method of tetraploid Robinia pseudoacacia, in particular to a method for tissue culture rapid propagation and gene transformation of tetraploid Robinia pseudoacacia, and belongs to the field of plant genetic engineering or the field of agricultural biotechnology. (2) Background technology [0002] Due to the lack of salt-resistant and drought-tolerant plant materials, especially the lack of arbor species, it has seriously restricted the construction of ecological projects and the development of rural economy in northern saline-alkaline arid areas. Tetraploid Robinia pseudoacacia is a new variety of Robinia pseudoacacia bred by chromosome doubling in Korea. In addition to maintaining the excellent characteristics of ordinary Robinia pseudoacacia (strong stress resistance, suitable for various climate and soil conditions, hard wood, wide use, flowers are excellent honey plants, There are rhizobia in the root, which...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01H4/00C12N5/14C12N15/53C12N15/82
Inventor 夏阳梁慧敏孙仲序王太明陈受宜
Owner SHANDONG FOREST SCI RES INST
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