Soybean MADS-box gene and applications thereof in floral organ modification

A soybean and gene technology, applied in the field of plant genetic engineering, can solve the problems of flower development research lag

Inactive Publication Date: 2010-08-18
NANJING AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] Soybean is an important grain and oil crop, and rese...

Method used

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  • Soybean MADS-box gene and applications thereof in floral organ modification
  • Soybean MADS-box gene and applications thereof in floral organ modification
  • Soybean MADS-box gene and applications thereof in floral organ modification

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Embodiment 1, cDNA cloning and identification of soybean MADS-box gene GmMADS1

[0037] Primers were designed according to the full-length sequence of GmMADS1,

[0038] Upstream primer: 5′-GAGATGGGAAGGGGAAGAGT-3′

[0039] Downstream primer: 5′-CAAGGAAGAGGCTAGCTAGG-3′

[0040]Using the RT-PCR method, the GmM Shuchang ADS1 gene was cloned from the soybean variety Jackson (publicly known and available for purchase from the National Soybean Improvement Center). Take the mixture of soybean flowers and flower buds, grind it with a mortar, add it to a 1.5mL EP tube filled with lysate, shake it fully, and then transfer it into a glass homogenizer. After homogenization, transfer to 1.5mL EP tube, and extract total RNA (TRIzol Reagents, Invitrogen, USA). The quality of total RNA was identified by formaldehyde denaturing gel electrophoresis, and then the RNA content was determined on a spectrophotometer. The obtained total RNA was used as a template, and reverse transcription ...

Embodiment 2

[0041] Embodiment 2, expression characteristics of soybean GmMADS1 gene in different tissues

[0042] The soybean variety Jackson was planted in the net room of Nanjing Agricultural University in normal season. Take the roots, stems and leaves of soybean at the six compound leaf stage, the mixture of flowers and flower buds at the full flowering stage (R1), the pods at the initial pod stage (R3), and the tender seeds 20 days after flowering, and store them in liquid nitrogen. Standby at -80°C. The whole soybean flower was aseptically divided into sepals, petals, stamens, and pistils at the full flowering stage, and was quickly frozen in liquid nitrogen and stored at -80°C for later use.

[0043] The extraction of total RNA was the same as in Example 1. Using soybean constitutively expressed Actin gene (GenBank accession No. V00450) as an internal reference, and using total RNA from different soybean tissues as templates, RT-PCR analysis was performed. RT-PCR analysis showed...

Embodiment 3

[0044] The genetic engineering application of embodiment 3, GmMADS1

[0045] According to the cDNA sequence of GmMADS1, primers were designed to amplify the complete coding reading frame, and a BamH I restriction endonuclease site was introduced on the upstream primer, and a SacI restriction endonuclease site was introduced on the downstream primer:

[0046] Upstream primer: 5-A GGATCC GAGATGGGAAGGGGAAGAGT-3

[0047] Downstream primer: 5-A GAGCTC CAAGGAAGAGGCTAGCTAGG-3

[0048] Using the PCR amplification product obtained in 1) as a template, after PCR amplification, the cDNA of GmMADS1 was cloned into the intermediate vector pMD18-T, and further cloned into the binary expression vector pBI121; the obtained expression vector was transformed into Agrobacterium strain LBA4404 was introduced into tobacco, and tobacco was transformed by Agrobacterium-mediated method. The obtained transgenic plants were analyzed by phenotype and cytology. It was found that the ectopic expressio...

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Abstract

The invention discloses a soybean MADS-box gene and applications thereof in floral organ modification, belonging to the field of biotechnology. GmMADS1, transcription factor in nucleus, is expressed in all the floral organs of four circles and, however, expressed in large quantity in petals and stamens, tobacco expressing excessive GmMADS1 increases the number of sepals, stamens and petals, a transformation from the sepals and the stamens to the petals is formed, as a result, the occurrence of cracking petals and bending stamens etc. causes the stamens to fail to come into contact with pistils, thus normal pollination and normal anther dehiscence cannot be realized and pollen activity is lowered. The above results represent that GmMADS1 plays an important regulation and control role in determining the position and the number of the petals and the development of male organs. New floral organ-variant material, resulted from the GmMADS1 gene, can be used for the breeding of crops and ornamental horticulture plants.

Description

technical field [0001] The invention relates to the application of a soybean MADS-box gene GmMADS1, which belongs to the field of plant genetic engineering. Background technique [0002] People have isolated a large number of MADS-box genes from the entire huge group of plants at different evolutionary positions. MADS-box genes play an important role in flower development, whether it is the transition from vegetative growth to reproductive growth; Also in the initial stage of reproductive growth, floral meristem-determining genes initiate the development of floral meristem; and in the later stage of reproductive growth, they play an important role in the development of floral organ primordia. [0003] Soybean is an important grain and oil crop, and the research on its flower development is relatively lagging behind. Soybeans have many flowers, small flower vessels, and heavy pollen. When the flowers are open, the wing petals and keel petals tightly cover the stamens and sti...

Claims

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Application Information

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IPC IPC(8): C12N15/29C12N15/10C12N15/82A01H5/00
Inventor 黄方喻德跃迟英俊
Owner NANJING AGRICULTURAL UNIVERSITY
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