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Corn saline-alkaline tolerance related molecular marker developed based on transcription factor gene ZmNAC89 and application thereof

A technology of molecular markers and transcription factors, applied in the direction of recombinant DNA technology, DNA/RNA fragments, microbial determination/inspection, etc., can solve problems affecting production

Pending Publication Date: 2021-03-30
NORTHEAST AGRICULTURAL UNIVERSITY
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0002] Salt-alkali stress is one of the important abiotic stress factors affecting the growth and development of maize, which seriously affects the yield

Method used

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  • Corn saline-alkaline tolerance related molecular marker developed based on transcription factor gene ZmNAC89 and application thereof
  • Corn saline-alkaline tolerance related molecular marker developed based on transcription factor gene ZmNAC89 and application thereof
  • Corn saline-alkaline tolerance related molecular marker developed based on transcription factor gene ZmNAC89 and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] Example 1. Sequence Variation Analysis of Maize Transcription Factor Gene ZmNAC89

[0026] According to the MaizeGDB database (https: / / www.maizegdb.org / ) to provide the ZmNAC89 gene sequence in the maize B73 genome, primers were designed segmentally, and the gene sequence and promoter sequence were amplified from 140 maize inbred lines. The ZmNAC89 gene primers are 89-1F / R and 89-2F / R, and the promoter primer is 89-p-F / R. The primer sequences are shown in Table 1.

[0027] Table 1 ZmNAC89 gene sequence and promoter sequence cloning primers

[0028]

[0029] The sequencing results were spliced ​​into a complete sequence using DNAMAN software, and multiple sequences were compared. The ZmNAC89 gene sequence was analyzed using DnaSPv6.0 software, and divided into different haplotypes based on the difference in SNP sites. D, Fu and Li's D* and Fu and Li's F* and other three methods were used for neutral test to preliminarily determine excellent haplotypes.

[0030] 1. H...

Embodiment 2

[0039] Example 2 Association Analysis of Genetic Variation of ZmNAC89 Gene and Saline-Alkaline Tolerance

[0040] TASSEL 5.0 software was used to screen out the variant sites with a typing rate MAF>0.05, and based on the phenotype data of the natural population’s salt-alkaline tolerance identification in the previous period, the association analysis between markers and phenotypes was carried out. The general linear model GLM was used to analyze the correlation between the 16 polymorphic sites in the ZmNAC89 gene coding region sequence and the salt-alkaline tolerance phenotype indicators of the tested maize inbred lines.

[0041] The results showed that at the 0.05 level, there were 5 non-synonymous mutation SNP sites associated with the alkali resistance phenotype index (see Table 4 and Table 5). In the basic analysis, the four SNP loci, including snp124, snp145, snp253 and snp375, were significantly correlated with plant height (SL) at the same time, all of which were non-syn...

Embodiment 3

[0047] Example 3 Development of dCAPS functional markers

[0048] According to the results of association analysis of ZmNAC89 gene coding region, dCAPs markers were developed at snp253(T / C) and snp767(G / C) loci. Since the primers were designed based on the reverse complementary strand of the reference sequence, snp253 was transformed from T / C to A / G mutation. Utilize the online website dCAPs Finder to analyze the best enzyme cutting sites and select restriction endonucleases, develop functional markers, dCAPs marker primers and corresponding endonucleases. dCAPs labeled primers and corresponding endonucleases are shown in Table 6.

[0049] The snp253(T / C) transformed marker is named DNdCAPS253, the PCR amplified fragment is 558bp in length, the 180th base is the mutation site, and the salt-alkali-sensitive material is T, which cannot be cut by the SacII restriction endonuclease; The saline-alkaline material is C, which can be cut into two fragments of 179bp and 379bp by SacI...

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Abstract

The invention discloses a corn saline-alkali tolerance related molecular marker developed based on a transcription factor gene ZmNAC89 and application thereof The molecular marker DNdCAPS253 is developed by analyzing sequence variation of a coding region and a promoter of an NAC transcription factor gene ZmNAC89 induced and expressed by saline-alkali stress in different maize inbred lines, performing correlation analysis on the sequence variation and saline-alkali resistance of the inbred lines and excavating correlation sites, contains SNP sites, adopts a saline-alkali sensitive material T, and cannot be cut by SacII restriction endonuclease; and the saline-alkaline tolerant material is C and can be cut into two fragments by SacII. Furthermore, the invention also provides an application of the molecular marker in corn saline-alkaline tolerance molecular marker-assisted breeding, the average detection efficiency of the molecular marker in identification of a saline-alkaline sensitive inbred line is 85.65%, and the invention provides a new technical means for screening saline-alkaline tolerance corn materials.

Description

technical field [0001] The present invention relates to a molecular marker related to maize salt-alkali tolerance and its application, in particular to a molecular marker related to maize saline-alkali tolerance developed based on the transcription factor gene ZmNAC89 and its use in screening maize germplasm resources for saline-alkali tolerance Applications. The invention belongs to the technical field of crop breeding. Background technique [0002] Salinity-alkali stress is one of the important abiotic stress factors affecting the growth and development of maize, which seriously affects the yield. Excavating salt-alkali tolerance-related genes and excellent germplasm resources is crucial for breeding salt-alkali-tolerant maize varieties. [0003] The genes related to salt-alkali tolerance that have been discovered so far mainly include genes related to proline synthesis, genes related to betaine synthesis, genes related to sugar alcohol synthesis, genes related to protec...

Claims

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Application Information

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IPC IPC(8): C12Q1/6895C12N15/11
CPCC12Q1/6895C12Q2600/13C12Q2600/156
Inventor 邸宏张林李春翔孙晓慧周羽封陈晨王振华曾兴
Owner NORTHEAST AGRICULTURAL UNIVERSITY
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