Maize transcription factor zmbhlh2 and its application

A technology of transcription factor and maize, applied in the field of genetic engineering, can solve the problem that the function of maize bHLH transcription factor is poorly understood, and achieve the effects of improving plant drought resistance, improving tolerance and small environmental impact

Active Publication Date: 2018-02-16
NORTHEAST AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

But little is known about the function of maize bHLH transcription factors in stress response

Method used

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  • Maize transcription factor zmbhlh2 and its application
  • Maize transcription factor zmbhlh2 and its application
  • Maize transcription factor zmbhlh2 and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] Cloning of Example 1 Maize Transcription Factor ZmbHLH2 Gene

[0040] The maize transcription factor ZmbHLH2 gene was cloned from the leaves of the maize variety Dongnong 255 (this maize variety has been approved and promoted by relevant departments) cultivated by the College of Agriculture of Northeast Agricultural University by RT-PCR technology. The specific method is as follows:

[0041] 1. Extract the total RNA from the maize leaves, and reverse transcribe to obtain the first strand of cDNA.

[0042] 2. The cDNA sequence of the transcription factor ZmbHLH2 gene was obtained by PCR amplification reaction using the first strand of DNA as a template and ZmbHLH2F and ZmbHLH2R as primers.

[0043] Wherein, the sequences of primers ZmbHLH2F and ZmbHLH2R described in step (2) are as follows:

[0044] ZmbHLH2F: 5′-ATCCGCATTCTGAACCATT-3′

[0045] ZmbHLH2R: 5'-TGCCTATCGTGTTGTAACC-3'.

[0046] Use 2×Taq PCR MasterMix (Tiangen Biochemical) for PCR amplification, the reactio...

Embodiment 2

[0049] Expression analysis of embodiment 2 ZmbHLH2 gene when subjected to stresses such as drought

[0050] The expression pattern of ZmbHLH2 gene in different drought-resistant inbred lines was detected by fluorescent quantitative PCR. Drought-resistant inbred lines, Zheng 58, Ji 853, Qi 319, Han 21, X178, 7922, Ye 52106 and K12; drought-sensitive inbred lines, Dong 91, Dong 46, Dong 26-6, Dong 34, red corn and PH4CV (See figure 2 ), the expression of ZmbHLH in drought-resistant inbred lines was significantly higher than that in non-drought-resistant inbred lines.

[0051] The following fluorescent quantitative PCR detection primers were designed according to the ZmbHLH2 gene sequence:

[0052] ZmbHLH2-QF1: 5′-GACGCCATCCTCCTACATC-3′

[0053] ZmbHLH2-QR1: 5′-GCTCTCCTCCTTGATTACC-3′

[0054] ZmACTIN-F: 5′-ACCTCACCGACCACCTAATG-3′

[0055] ZmACTIN-R: 5′-GCAGTCTCCAGCTCCTGTTC-3′

[0056] ZmACTIN (GenBank accession number: GRMZM2G126010) was used as an internal reference gene....

Embodiment 3

[0058] Example 3 Obtaining and Drought Resistance Verification of ZmbHLH2 Gene-transferred Arabidopsis Plants

[0059] Total RNA was extracted from maize leaves, and the first strand of cDNA was obtained by reverse transcription. The cDNA sequence of the transcription factor ZmbHLH2 gene was obtained by PCR amplification reaction using the first strand of DNA as a template and ZmbHLH2F and ZmbHLH2R as primers, and its cDNA fragment was ligated into the pMD18-T vector and transformed into Escherichia coli DH5α strain for cloning and Sequencing verification, the correct plasmid was named ZmbHLH2-18T; using the ZmbHLH2-18T plasmid as a template, ZmbHLH2-ORF-F: 5′-CG GAATTC ATGAACCTGTGGACG-3' (the underline is the EcoR I restriction site) and ZmbHLH2-ORF-R:5'-CG GGATCC CCTGCCCATGGCAGA-3' (the underline is the BamHI restriction site) is used as a primer to amplify the ORF of the ZmbHLH2 gene, and the amplified product is double-digested by EcoR I and BamHI and then connected to t...

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Abstract

The invention relates to a corn transcription factor ZmbHLH2 and application thereof. The nucleotide sequence of the corn transcription factor ZmbHLH2 is shown as the SEQ ID NO.1. The transcription factor is related to drought resisting of corn. By conducting drought resisting function verification of the gene on ZmbHLH2 gene-modified arabidopsis thaliana, it is proved that the endurance capacity of the ZmbHLH2 gene-modified arabidopsis thaliana to drought stress is improved. The gene can be applied to drought-resisting transgene study on the corn and other crops. The corn transcription factor ZmbHLH2 comes from the corn, gene flow cannot be generated, and a new gene resource is provided for improving the drought resistance of plants.

Description

technical field [0001] The invention relates to the technical field of genetic engineering, in particular to the maize transcription factor ZmbHLH2 gene and its application. Background technique [0002] Maize cultivation is often threatened by drought, which affects maize photosynthetic and physiological activities, grain formation and yield. In view of the important role of maize in food security, the drought resistance of maize has been studied from the aspects of population genetic improvement, cultivation measures and physiological and biochemical levels. At the population level, nine drought-resistant inbred lines were screened by studying the total root length (TRL) and root surface area (TSA) of maize inbred lines closely related to drought resistance. A study of 13 drought-resistant populations found that there was a significant positive correlation between palmitic acid and oleic acid in drought-resistant maize varieties. At the cultivation level, it was found th...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29C12N15/82A01H5/00A01H6/20A01H6/46C12Q1/6895C12N15/11
CPCC07K14/415C12N15/8205C12N15/8273C12Q1/6895C12Q2600/158
Inventor 董玲周羽刘显君曾兴
Owner NORTHEAST AGRICULTURAL UNIVERSITY
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