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Tobacco slowly activating anion channel protein NtSLAH8 and application thereof

A channel protein and anion technology, applied in the field of tobacco slow anion channel protein NtSLAH8 and its application patent application, which can solve problems such as large functional differences

Inactive Publication Date: 2019-02-15
ZHENGZHOU TOBACCO RES INST OF CNTC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

but due to SLAC There are many genes in the gene family, and their functions vary greatly in different plants. Therefore, for different plants and different SLAC The function of the gene still needs to be further studied and differentiated, so that it can be targeted for future plant improvement

Method used

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  • Tobacco slowly activating anion channel protein NtSLAH8 and application thereof
  • Tobacco slowly activating anion channel protein NtSLAH8 and application thereof
  • Tobacco slowly activating anion channel protein NtSLAH8 and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0047] Present embodiment mainly with regard to tobacco slow anion channel protein NtSLAH8 The process of gene acquisition is briefly introduced as follows.

[0048] Using the cultivar tobacco leaf as a sample, the total RNA of the tobacco leaf was extracted with an RNA extraction kit, and reverse-transcribed into cDNA for future use;

[0049] The sequences of the amplification primers were designed as follows:

[0050] F: 5'-CGCGAGCTCGGTACCATGGTTGCAAATCCAACT-3',

[0051] R: 5'-GCTCACCATGGATCCATTACGTTTAGTGAAGT-3';

[0052] Using the prepared cDNA as a template, PCR amplification was performed using the above primers to obtain the sequence of the NtSLAH8 gene.

[0053] The PCR reaction system is:

[0054] Upstream primer 1 μL,

[0055] Downstream primer 1 μL,

[0056] cDNA 1 μL,

[0057] 10×buffer 5μL,

[0058] dNTPs 6 μL,

[0059] EazyTaq enzyme 1 μL,

[0060] ddH 2 0 to 50 μL.

[0061] PCR reaction program: pre-denaturation at 94°C for 5 min; denaturation at 94°C...

Embodiment 2

[0070] to be sure NtSLAH8 Gene function in tobacco, selection NtSLAH8 The specific nucleic acid fragment in the gene (the 1st-388th nucleotide sequence of SEQID NO.1 in the sequence table) is used as a guide sequence to construct a silencer NtSLAH8 The VIGS vector was used for the transient silencing of the gene, and the tobacco plants were further transformed to construct transgenic plants. The relevant experimental procedures are briefly introduced as follows.

[0071] (1) Construction of VIGS vector for transient silencing

[0072] First, the primer sequences for PCR amplification were designed as follows:

[0073] NtSLAH8 -F:5'-TCGACGACAAGACCCTGCAGATGGTTGCAAAATCCAAC-3',

[0074] NtSLAH8 -R: 5'-TGAGGAGAAGAGCCCTGCAGGTGGAAATGAGTAAGAC-3';

[0075] Perform PCR amplification with the above primer sequences (amplification length: 388bp) to obtain the guide sequence of VIGS;

[0076] Secondly, using the In-Fusion method, the above-mentioned amplified guide sequence was...

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Abstract

The invention belongs to the technical field of tobacco genetic engineering, and particularly relates to tobacco slowly activating anion channel protein NtSLAH8 and application thereof. A gene CDS sequence comprises 579bp basic group, with the base sequence being as shown in SEQ ID NO.1; the first to 388th nucleotide for a specific nucleic acid fragment. The tobacco slowly activating anion channelprotein NtSLAH8 belongs to key protein for metabolism of tobacco chlorine ion, and the chlorine ion content in transgenic silencing plant is obviously reduced after NtSLAH8 gene expression is inhibited through a virus induced gene silencing (VIGS) technology, so that a good foundation can be laid for cultivation of low chlorine content novel varieties, and meanwhile a fundamental technical support for the stabilizing of tobacco leaf quality and improvement of cigarette quality.

Description

technical field [0001] The invention belongs to the technical field of tobacco genetic engineering, and specifically relates to a tobacco slow anion channel protein NtSLAH8 and a patent application for its application. Background technique [0002] Existing studies on tobacco generally believe that the appropriate content of chloride ions in tobacco leaves is 0.3-0.8, and when the content reaches 1%, it will affect the smoldering fire retention, and when it is further higher than 1%, there will be black ash flameout phenomenon; On the one hand, when the content of chloride ions in tobacco leaves is too high, starch will accumulate more, the leaves will be thick and brittle, and have high hygroscopicity, which will make the color darken easily during storage and produce bad smell. In a word, the chloride ion content in tobacco leaves has a relatively important and direct impact on the quality of tobacco leaves. [0003] Based on the direct impact and importance of chloride i...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/29C12N15/10C12N15/82C07K14/415A01H5/00A01H6/82
CPCC07K14/415C12N15/8218C12N15/8243
Inventor 张慧徐国云卢鹏郑庆霞金静静翟妞金立锋周会娜许亚龙刘萍萍陈千思李泽锋
Owner ZHENGZHOU TOBACCO RES INST OF CNTC
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