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Tomato sllcd gene and its application

A tomato and gene technology, applied in the field of genetic engineering, can solve problems such as unknown functions, and achieve the effect of maintaining excellent traits and good application value

Active Publication Date: 2020-07-10
SHANXI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Currently, other endogenous H 2 S production-related enzyme genes, the functions of tomato fruit ripening and seed development are still unknown

Method used

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  • Tomato sllcd gene and its application
  • Tomato sllcd gene and its application
  • Tomato sllcd gene and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] Cloning of embodiment 1 tomato SLLCD coding region gene

[0032] Utilize the forward primer of nucleotide sequence as shown in SEQ ID No.3 and the reverse primer of nucleotide sequence as shown in SEQ ID No.4, clone the coding region sequence of SLLCD gene from the carpel cDNA of tomato MT ;

[0033] PCR program: 94°C, 5 minutes; 94°C, 30 seconds; 55°C, 30 seconds; 72°C, 1 minute 30 seconds; repeat 35 times; 72°C, 10 minutes.

[0034]

[0035] The above-mentioned PCR product was connected to the cloning vector pET28a by the method of double digestion (such as figure 2shown); the ligation product was transformed into Escherichia coli DH5α, and multiplied therein, and the positive clone was sequenced and screened to obtain the cDNA sequence of the coding region of tomato SLLCD gene, its nucleotide sequence is shown in SEQ ID No.1, encoded by it The amino acid sequence of the protein is shown in SEQ ID No.2.

Embodiment 2

[0036] Example 2 Sequence Structure Analysis of Tomato SLLCD Protein

[0037] Tomato SLLCD has the ability to produce endogenous H 2 The function of S has 63% similarity with the protein sequence of LCD coding region in Arabidopsis. The result is as figure 1 shown.

Embodiment 3

[0038] Embodiment 3 tomato SLLCD gene expression vector and transgenic plant

[0039] tomato endogenous H 2 S produces a section of 3' end sequence (400bp, its nucleotide sequence as shown in SEQID No.5) of enzyme-related gene SLLCD, reversely constructed in Figure 4 The shown intermediate vector pART27 is used to inhibit the expression level of SLLCD in tomato by using the principle of RNA interference, and then to study its function.

[0040] Through the Agrobacterium-mediated transformation method, the 3' end sequence of the SLLCD gene cDNA carried by pART27 was transferred into tomato, and the cotyledons after aseptic sowing of tomato seeds were used as recipients to obtain transformed tomato plants, and the selection marker in the plants was kana Mycin. Agrobacterium-mediated transformation methods and steps are as follows:

[0041] The method and steps of Agrobacterium transformation tomato are as follows:

[0042] 1. Seed Preparation

[0043] Wash 1.5g of tomato s...

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Abstract

The invention provides a tomato SLLCD coding region gene, and tomato SLLCD protein encoded by the gene; a tomato coding region nucleotide sequence of the tomato SLLCD gene is shown as in SEQ ID No. 1. The tomato SLLCD gene and the application thereof have the advantages that the tomato LSSCD coding region gene is an enzyme coding gene related directly to the production of endogenous H2S, can inhibit SLLCD gene expression in tomato to significantly affect the development of seeds in tomato, and transgenic plants having zero seeds or incompletely developed seeds; the gene is applicable to the acquisition of seedless tomato and maintenance of beneficial matters in fruits; in addition, growth of fruits on SLLCD-transgenic plants is not affected; therefore, good traits of tomato can be maintained; it is indicated that the SLLCD gene is well applicable in the development process.

Description

technical field [0001] The invention relates to the field of genetic engineering, in particular to a tomato endogenous H 2 S produces directly related enzyme-encoding genes and their application in the process of producing seedless tomato. Background technique [0002] Seed formation is an important process in plant development. Sexually reproduced plants develop from ovules through the process of pollination and fertilization. At the same time, the reproduction mode and genetic variation characteristics of plants can be used to maintain their excellent varieties. Seedlessness is an excellent fruit trait. Seedless fruit is of good quality and high edible rate, which is more likely to attract consumers to buy. The production of seedless fruit can improve fruit quality, achieve a higher seedless rate, increase the size of fruit grains, thereby maintaining the beneficial substances in the fruit to the greatest extent, obtain excellent vegetable and fruit varieties, increase ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/52C12N9/00C12N15/82A01H5/00A01H6/82
CPCC12N9/00C12N15/8287
Inventor 裴雁曦张彦洁刘旦梅
Owner SHANXI UNIV
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