Method for sequencing clone genes with recessive mixed pools

Abstract

The invention discloses a method for sequencing clone genes with recessive mixed pools. The method comprises the following steps of constructing segregation populations by hybridizing parents; allocating pools for the segregation populations according to individual target properties; obtaining dominant and the recessive specific loci of the parents through high-throughput sequencing; comparing with the sequencing results of the recessive mixed pools to obtain candidate genes; gathering the candidate genes in the recessive mixed pools and re-sequencing clone target genes; The method is on the basis of sequencing, can be used for any species, and directly clones genes themselves. With the method, workload is decreased greatly, speed is substantially accelerated; and risks are reduced.

Description

technical field [0001] The invention belongs to the field of genetics and discloses a recessive mixed pool sequencing gene cloning method. Background technique [0002] Species in nature have rich and diverse traits, such as plant height, disease resistance, yield, etc. Since Mendel's time, it has been gradually realized that traits are controlled by genetic factors. Morgan further defined genetic factors as "genes", and pointed out that genes are located on chromosomes and arranged linearly. Determining the specific position of a gene on a chromosome (gene mapping) is the basis for studying traits and the prerequisite for isolating, cloning and utilizing genes. [0003] The classic gene localization strategy is derived from Morgan's linkage theory, that is, adjacent genes on chromosomes (ie, gene linkage) cannot segregate freely, but tend to be passed on to offspring as a whole, and the traits they control also tend to appear at the same time. The closer the two are, the ...

AI Technical Summary

Problems solved by technology

However, only a small portion of species have developed such markers, and most of the beneficial genes in nature cannot be located, cloned and utilized by this method

[0006] (2) What is found is the molecular marker linked to the target gene, that is, the segment containing the target gene is obtained instead of the target gene itself, and gene prediction and a lot of verification work are required to exclude non-target genes in the segment. When the segment is large, the work is very difficult or even impossible to complete

causing some problems

First, excessive genetic distance often leads to partial segregation of traits, and it is necessary to establish a separate population to investigate the trait segregation ratio, which increases the workload

Second, distant hybridization is difficult to use in breeding, which leads to a disconnect between the theoretical research of gene mapping and the practice of breeding application, which violates the original intention of gene mapping

[0008] (4) During fine mapping, the distance between the linkage marker and the target gene is very close, and it becomes more and more difficult to find polymorphic markers, and there are often cases where no marker is available

However, the exchange rate calculation is a quantitative analysis, which needs to check every single plant in the population, and the workload is very huge.

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