Regulatory gene of rice dwarf brittle mutant dbc1 and its application

A technology for regulating genes and mutants, which is applied in the field of genetic engineering, can solve the difficult problems of straw treatment, etc., and achieve the effect of regulating rice cell division and promoting breeding research

Active Publication Date: 2021-09-28
SOUTHWEST UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Due to the toughness of straw, it is not easy to crush and return to the field, and straw disposal has always been a thorny issue

Method used

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  • Regulatory gene of rice dwarf brittle mutant dbc1 and its application
  • Regulatory gene of rice dwarf brittle mutant dbc1 and its application
  • Regulatory gene of rice dwarf brittle mutant dbc1 and its application

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0060] Implementation 1: Cloning, sequencing and functional verification of rice dwarf brittle mutant gene DBC1

[0061] 1. Cloning and sequencing of rice dwarf brittle gene DBC1

[0062] On the basis of fine mapping of the mutant gene DBC1 in the previous stage, the present invention uses online gene prediction (http: / / mendel.cs.rhul.ac.uk), BLAST online comparison (http: / / blast.ncbi.nlm.nih .gov / ) and functional analysis of gene overexpression. Through map-based cloning and sequence comparison, it was found that the gene was due to the mutation of the 5th base of the 5th intron of the gene LOC_Os02g50550 from the normal C to T. Since genes need to be modified, spliced ​​and spliced ​​after transcription, that is, intron mutations may affect the splicing and editing of precursor messenger RNA. Therefore, design a pair of primers P1 and P2 with a distance of about 550bp on the cDNA, including the mutation site, see image 3 B, where the gray box represents the exon, the bla...

Embodiment 2

[0065] Example 2: Experimental analysis of the hormone response of the rice dwarf brittle mutant gene DBC1

[0066] 1) Germinate the mutant and wild-type seeds, and select the seeds with the same dew and whiteness to sow in the seedlings containing 0 and 10 -10 、10 -8 、10 -6 and 10 -4 μM gibberellin (GA) nutrient solution, placed in a constant temperature incubator for cultivation. Add equal amount of nutrient solution and nutrient solution containing gibberellin in corresponding concentration every day. The length change of the second leaf sheath was measured and counted every 24 hours until the stage of two leaves and one heart. And at the one-leaf-one-heart stage (about the 7th day), the leaves under each treatment were taken, RNA was extracted, reverse-transcribed into cDNA, and stored for future use. It was found that when the gibberellin concentration was lower than 10 -8 At μM, the growth of the second leaf sheath of the wild type and the mutant was almost indisti...

Embodiment 3

[0071] Example 3: Cell division speed experiment of mutant DBC1

[0072] Paraffin sections were used to analyze the cells at the growth point of the root tip. Under the same cell number, the average number of binucleated monocytic cells in DBC1 was 7.4, and it was 4.1 in the wild type, showing a significant level. See Figure 7 A-E. This implies that the mutant cells divide at a slower rate. RT-PCR analysis of the expression changes of cyclin-related genes in wild type, mutants and overexpression lines OE-1, RT-PCR quantitative primer sequences are shown in Table 3, the results show that the expression of cyclin-related genes in mutants Compared with the wild type, the expression of CycB, CycD, CycP and CycT was restored in the transgenic lines, see Figure 7 F. This further explains the slowed cell division in the mutant DBC1. That is, mutations in this gene cause the rate at which cells divide to slow down.

[0073] Table 3 cyclin RT-PCR quantitative primer sequence

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Abstract

The invention belongs to the technical field of genetic engineering, and in particular relates to a regulation gene of rice dwarf brittle mutant DBC1 and its application. By cloning and sequencing the rice dwarf brittle mutant DBC1, the present invention excavates the regulation gene of rice dwarf brittle stalk, and experimentally verifies that the gene has the function of regulating rice dwarf brittle stalk, providing a basis for rice transgenic research It is a powerful tool to promote the breeding research of high-yield and high-quality rice.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, and in particular relates to a regulation gene of rice dwarf brittle mutant DBC1 and its application. Background technique [0002] Rice is the most important food crop in my country, and more than half of the population takes it as a staple food; at the same time, because of its small genome and high homology, rice is the earliest cereal crop sequenced, making it a monocot Model plants for plant molecular biology research. The "Green Revolution" and the application of hybrid rice have greatly increased the quantity of rice, which is of great significance to solving my country's food security. Straw returning to the field is a measure of fertilizing soil fertility and increasing production that is generally valued in the world today. It can increase fertilizer and increase production while eliminating the air pollution caused by straw burning. In addition, straw returning can increas...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29C07K14/415C12N15/82A01H5/04A01H6/46
CPCC07K14/415C12N15/8261
Inventor 桑贤春张晓琼史岭谢佳王晓雯何光华杨正林凌英华
Owner SOUTHWEST UNIV
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