High-throughput method for segregating quantitative character regulatory gene

Abstract

The invention discloses a high-throughput method for segregating a quantitative character regulatory gene, which comprises the following steps of: 1) construction of a target character segregation population, in which the population is a population in two parent hybridization progenies (F2 and F3), a DH system and a RIL; 2) mixing of extreme samples and segregation of total RNA in the population, in which a progeny segregation population is divided into three categories according to character phenotype; 3) gene expression analysis, in which the difference and sameness of gene expressions between two extreme mixed samples are compared by utilizing a gene expression analysis method, namely one of chip, EST sequencing, subtraction, cDNA-AFLP, and the like; and 4) acquisition and verification of a candidate gene, in which a differential expression gene between the two extreme mixed samples is found and is a candidate regulatory gene related to target character, and the function of the gene is verified through transgene, gene expression, molecular marker correlation and a contribution rate analysis method to obtain a target gene with regulatory character phenotype. The method is suitable for the segregation of a certain quantitative character regulatory gene controlled by multigene of all organisms, and is a simple, quick, high-throughput and economical gene segregation method.

Description

technical field [0001] The invention belongs to the field of biological technology, and in particular relates to a high-throughput method for isolating quantitative trait regulation genes. It is suitable for all kinds of organisms such as plants and animals that can obtain offspring through hybridization, and is especially suitable for the isolation of important agronomic trait genes of various crops. Background technique [0002] Biological traits are controlled by genes, and traits can be divided into qualitative traits and quantitative traits. Quality traits are characterized by discontinuous distribution of individual traits in segregated populations, and quality trait genes are also called main effect genes. Because quality trait genes significantly control phenotypes, and for a certain quality trait, there are few control genes, so the cloning method is relatively simple, including gene mapping map-based cloning, mutant analysis, and isolation of differentially expres...

AI Technical Summary

Problems solved by technology

[0011] The disadvantages of this method are: long experimental cycle, cumbersome steps, complicated operation, heavy workload, high consumption, and high false positives

This method has the following deficiencies: 1) only functional genes with large effects can be cloned, and the gene loci with small effects cannot be screened out for corresponding near-isogenic lines because of small differences in traits; There are often multiple minor functional genes that are jointly affected, so this method has certain unknowns; 2) The experimental cycle is long, and it takes about 5 years to construct a near-isogenic line; Each regulatory gene of a quantitative trait needs to carry out similar repeated research work, and the quantitative trait is controlled by multiple genes. To separate multiple genes, the work cost and workload will be doubled several times.

One of its biggest disadvantages is the huge investment in manpower and material resources. Large-scale gene expression analysis must be carried out for each individual in the group. The scope of application is very narrow, and research can only be carried out on certain model crops. This is why this technology has not been widely promoted. s reason

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