Method for sequencing clone genes with parent specific loci

Abstract

The invention discloses a method for sequencing clone genes with parent specific loci. The method comprises the following steps of selecting materials different target properties; constructing segregation populations by hybridizing two parents; constructing a gene pool by selecting individuals having recessive properties from the segregation populations; high-throughput sequencing the genomes of the two parents; obtaining dominant and the recessive specific loci of the parents through preliminary direct assembly and comparison, wherein the specific loci are candidate loci of the target genes; determining the target gene loci by detecting every candidate locus of each individual through gathering the candidate loci of the recessive pool and re-sequencing or a PCR mode; cloning full-length target genes via PCR amplification; and verifying functions of the target genes via genetic complementary experiments. 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 method for sequencing and cloning genes at specific sites of parents. 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 ...

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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