Application of Rapeseed bnaspt3 Gene in Promoting Dicotyledonous Silique Growth

Abstract

The invention discloses an application of a brassica napus BnaSPT3 gene in promoting growth of pods of dicotyledons. A base sequence of the brassica napus BnaSPT3 gene is shown as SEQ ID NO.1. The application comprises the following steps: (1) linking the brassica napus BnaSPT3 gene into a plant expression vector, so that a recombinant expression vector is constructed; and (2) transforming the recombinant expression vector into a recipient plant. The invention also discloses the recombinant expression vector containing the brassica napus BnaSPT3 gene and a transformant containing the recombinant expression vector. According to the application provided by the invention, by cloning the brassica napus BnaSPT3 gene, implementing genetic transformation in arabidopsis thaliana and through excessive expression of the exogenous brassica napus BnaSPT3 gene, the size of arabidopsis thaliana spt mutant pods can be obviously increased, and the number of seeds can be recovered. On the basis of genetic engineering technology, over-expression of the brassica napus BnaSPT3 gene in a receptor is achieved, and the size of the pods is changed; therefore, the invention is conducive to high-yield researches.

Description

technical field

[0001] The invention relates to the technical field of bioengineering, in particular to the application of rapeseed BnaSPT3 gene in promoting the growth of dicotyledons siliques. Background technique

[0002] Rapeseed is one of the main oil crops in my country. The planting and production of rapeseed directly affect the safety of oilseed food in my country. And in the world, the position of rapeseed in the oil crops is also the top priority. At present, the output of rapeseed in our country is still far from meeting the domestic demand, and increasing the output of rapeseed is an urgent task. Silique is an important part of rapeseed yield components, and silique and its related characters have direct or indirect effects on rapeseed yield. Rapeseed silique, as the fruiting organ of rapeseed, is not only an important storage organ, but also an important photosynthetic organ, with dual functions of "sink" and "source". The growth and development of siliques s...

Examples

Embodiment 1

[0032] Example 1 Construction of Rapeseed bHLH Transcription Factor BnaSPT3 Gene Overexpression Vector and Arabidopsis Genetic Transformation

[0033] (1) Rapeseed (Brassica napus) leaves were taken, ground with liquid nitrogen, and RNAiso reagent (Takara Company) was added to extract RNA. 1 μg of RNA was taken for reverse transcription reaction to obtain cDNA.

[0034] (2) Design specific primers according to the gene sequence of BnaSPT3 (BnaA08g15580D) on Genbank, and add Sal I and Not I restriction enzyme sites to the 5' ends of the primers respectively. The sequences are as follows:

[0035] Primer 1: 5'-GCGTCGACATGGCTGATAAGAAATTGATTTCATCT-3';

[0036] Primer 2: 5'-TTGCGGCCGCTCAACTAAGTCGATCTTTTAGGTCAGGT-3'.

[0037] (3) Perform PCR amplification using the full-length cDNA as a template.

[0038] high fidelity enzyme HS DNA Polymerase (TaKaRa Code: DR010A) system:

[0039] 5×PrimeSTAR Buffer (Mg 2+ plus) 10 μL,

[0040] dNTP Mixture (2.5mM each) 4μL,

[0041] Prime...

Embodiment 2

[0059] The character observation of embodiment 2 Arabidopsis siliques

[0060] The siliques of Arabidopsis spt2 and spt12 mutants are shorter than those of the wild type, and are relatively flat in the middle near the top. We selected spt2 and spt12 mutants as transgene recipients.

[0061] Carry out silique length measurement record to the obtained transgenic Arabidopsis plant, such as figure 1 As shown, overexpression of the rape BnaSPT3 gene (sequence shown in SEQ ID NO.1) in Arabidopsis silique dysplasia mutants significantly increases the length of the mutant siliques and restores them to wild-type levels.

[0062] With the help of genetic engineering technology, the functional gene can be overexpressed in the recipient, and the size of the silique can be improved, which is helpful for high-yield research.

Embodiment 3

[0064] In order to detect whether the overexpression of the rapeseed gene BnaSPT3 can promote the growth of Arabidopsis wild-type siliques, we also conducted functional verification using the Arabidopsis wild-type plant Columbia as the transgenic receptor in Example 1.

[0065] The harvested transgenic Arabidopsis plants were measured and recorded for the length of siliques, and the statistical results were as follows: figure 2 shown. Overexpression of rapeseed BnaSPT3 gene in Arabidopsis wild-type can promote the growth of siliques, which are significantly larger than untransformed Arabidopsis wild-type plants.

[0066] Our experimental results took the model plant Arabidopsis as the background, and proved that the BnaSPT3 gene of rapeseed can promote the growth of dicotyledonous siliques.

[0067]

technical field

[0001] The invention relates to the technical field of bioengineering, in particular to the application of rapeseed BnaSPT3 gene in promoting the growth of dicotyledons siliques. Background technique

[0002] Rapeseed is one of the main oil crops in my country. The planting and production of rapeseed directly affect the safety of oilseed food in my country. And in the world, the position of rapeseed in the oil crops is also the top priority. At present, the output of rapeseed in our country is still far from meeting the domestic demand, and increasing the output of rapeseed is an urgent task. Silique is an important part of rapeseed yield components, and silique and its related characters have direct or indirect effects on rapeseed yield. Rapeseed silique, as the fruiting organ of rapeseed, is not only an important storage organ, but also an important photosynthetic organ, with dual functions of "sink" and "source". The growth and development of siliques s...

Examples

Embodiment 1

[0032] Example 1 Construction of Rapeseed bHLH Transcription Factor BnaSPT3 Gene Overexpression Vector and Arabidopsis Genetic Transformation

[0033] (1) Rapeseed (Brassica napus) leaves were taken, ground with liquid nitrogen, and RNAiso reagent (Takara Company) was added to extract RNA. 1 μg of RNA was taken for reverse transcription reaction to obtain cDNA.

[0034] (2) Design specific primers according to the gene sequence of BnaSPT3 (BnaA08g15580D) on Genbank, and add Sal I and Not I restriction enzyme sites to the 5' ends of the primers respectively. The sequences are as follows:

[0035] Primer 1: 5'-GCGTCGACATGGCTGATAAGAAATTGATTTCATCT-3';

[0036] Primer 2: 5'-TTGCGGCCGCTCAACTAAGTCGATCTTTTAGGTCAGGT-3'.

[0037] (3) Perform PCR amplification using the full-length cDNA as a template.

[0038] high fidelity enzyme HS DNA Polymerase (TaKaRa Code: DR010A) system:

[0039] 5×PrimeSTAR Buffer (Mg 2+ plus) 10 μL,

[0040] dNTP Mixture (2.5mM each) 4μL,

[0041] Prime...

Embodiment 2

[0059] The character observation of embodiment 2 Arabidopsis siliques

[0060] The siliques of Arabidopsis spt2 and spt12 mutants are shorter than those of the wild type, and are relatively flat in the middle near the top. We selected spt2 and spt12 mutants as transgene recipients.

[0061] Carry out silique length measurement record to the obtained transgenic Arabidopsis plant, such as figure 1 As shown, overexpression of the rape BnaSPT3 gene (sequence shown in SEQ ID NO.1) in Arabidopsis silique dysplasia mutants significantly increases the length of the mutant siliques and restores them to wild-type levels.

[0062] With the help of genetic engineering technology, the functional gene can be overexpressed in the recipient, and the size of the silique can be improved, which is helpful for high-yield research.

Embodiment 3

[0064] In order to detect whether the overexpression of the rapeseed gene BnaSPT3 can promote the growth of Arabidopsis wild-type siliques, we also conducted functional verification using the Arabidopsis wild-type plant Columbia as the transgenic receptor in Example 1.

[0065] The harvested transgenic Arabidopsis plants were measured and recorded for the length of siliques, and the statistical results were as follows: figure 2 shown. Overexpression of rapeseed BnaSPT3 gene in Arabidopsis wild-type can promote the growth of siliques, which are significantly larger than untransformed Arabidopsis wild-type plants.

[0066] Our experimental results took the model plant Arabidopsis as the background, and proved that the BnaSPT3 gene of rapeseed can promote the growth of dicotyledonous siliques.

[0067]

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