Corn waxy gene mutant as well as molecular marker and application thereof

A technology of molecular markers and protein mutants, applied in the fields of application, genetic engineering, plant gene improvement, etc., can solve problems such as easy loss, and achieve the effect of increasing the content of amylopectin and reducing the activity of granule-bound starch synthase

Active Publication Date: 2020-11-27
BEIJING ACADEMY OF AGRICULTURE & FORESTRY SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

In addition to the above five mutation types, there may still be unknown new waxy gene mutation types

Method used

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  • Corn waxy gene mutant as well as molecular marker and application thereof
  • Corn waxy gene mutant as well as molecular marker and application thereof
  • Corn waxy gene mutant as well as molecular marker and application thereof

Examples

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Effect test

Embodiment 1

[0041] The following examples are used to illustrate the present invention, but are not intended to limit the scope of the present invention. Example 1 Obtaining of maize waxy gene mutant wx-hAT

[0042] 200 germplasms of waxy maize inbred lines were collected from the maize germplasm resource bank of the Maize Research Center of Beijing Academy of Agriculture and Forestry Sciences, and the DNA of maize leaves was extracted using the plant DNA extraction kit of TianGen Company. The segment from exon 1 to exon 14 was amplified. Sequencing analysis revealed that among 200 germplasms of waxy maize inbred lines, 8 maize materials carried a new allelic mutation of the waxy gene. The first exon to the fourth exon of the waxy gene of these eight materials were amplified by PCR with E1-4F / R primers. PCR amplification conditions were: 94°C, 5min, then 94°C, 60s, 59°C, 60s, 72°C, 2min for 34 cycles, and finally 72°C extension for 10min. Electrophoresis detection found that the size of...

Embodiment 2

[0051] Example 2 cDNA sequence analysis of maize waxy gene mutant wx-hAT

[0052] The wx-hAT type waxy maize leaf RNA obtained in Example 1 was extracted by the Trizol method, and the RNA was reverse-transcribed into cDNA by a reverse transcription kit (Takara). The cDNA sequences were amplified with waxycDNAF2 and WaxycDNAR2 primers (Table 4). The PCR amplification conditions were: 94°C for 5min, then 38 cycles of 94°C, 60s, 56°C, 60s, 72°C, 2min, and finally 72°C for 10min. Electrophoresis detection found that the size of the amplified fragment in the common maize inbred line B73 was 163bp, while the amplified fragment in the wx-hAT type waxy maize HN2 was 1342bp ( figure 2 ). Sequencing analysis found that 1179 bp of the 2.2 kb insert was exonized in wx-hAT type waxy maize ( image 3 ), the exonized sequence contains a stop codon, resulting in premature termination of translation of the inserted protein. This result further confirms that the loss of function of the wax...

Embodiment 3

[0055] Example 3 Development and Application of Specific Functional Molecular Markers of Maize Waxy Gene Mutant wx-hAT

[0056] Based on the sequence difference between wx-hAT waxy maize and common maize in the third exon of the waxy gene, a site-specific PCR functional molecular marker PCR-hAT was developed. The amplification primer information of this molecular marker is shown in Table 5.

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Abstract

The invention relates to the technical field of plant molecular biology, in particular to a corn waxy gene mutant as well as a molecular marker and application thereof. The invention provides a corn waxy protein mutant, and compared with a wild WAXY protein, the protein mutant comprises mutation that an amino acid sequence represented by SEQ ID NO.1 is inserted behind the 125th amino acid resulting in early termination of protein translation, so that protein translation terminates early. The invention also provides a corn waxy gene mutant, and compared with a wild waxy gene, the gene mutant comprises mutation that a nucleotide sequence shown as SEQ ID NO.4 is inserted behind the 43rd nucleotide of the 3rd exon. According to the WAXY protein mutant and the waxy gene mutant provided by the invention, the amylopectin content of corn grains can be remarkably improved, and the grains have waxy characters. The invention further provides the molecular marker of the waxy gene mutant, seedlingstage selection of waxy corn can be achieved, and the breeding period is greatly shortened.

Description

technical field [0001] The invention relates to the technical field of plant molecular biology, in particular to a corn waxy gene mutant and its molecular marker and application. Background technique [0002] The grains of corn and other grains are rich in starch. The content and composition of starch (such as the content of amylose and amylopectin) are important factors that affect the nutritional value, processing quality and economic value of the grain. Waxy corn, also known as sticky corn, has high nutritional, processing and economic value. Waxy corn kernel endosperm contains close to 100% amylopectin, so it is sticky. Waxy maize is mainly consumed as food in Asia and is also an important raw material for the textile and paper industries. Waxy corn originated in China and was introduced to the United States by American Collins in 1908, and then spread all over the world. [0003] Waxyness in maize is controlled by a single recessive mutation in waxy, which encodes gr...

Claims

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Application Information

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IPC IPC(8): C12N9/10C12N15/54C12N15/11C12N15/82A01H5/00A01H6/46C12Q1/6895
CPCC12N9/1051C12Y204/01021C12N15/8245C12Q1/6895C12Q2600/13C12Q2600/156
Inventor 骆美洁赵久然赵衍鑫卢柏山史亚兴张云霞
Owner BEIJING ACADEMY OF AGRICULTURE & FORESTRY SCIENCES
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