Method and device for detecting cytoplasmic inheritance

A cytoplasmic and sample technology, applied in the field of cytoplasmic genetic detection, can solve problems such as misjudgment, and achieve high resolution, high sensitivity, and flexible sampling.

Active Publication Date: 2022-05-10
ZHEJIANG ACADEMY OF AGRICULTURE SCIENCES
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, due to the multi-copy nature of cytoplasmic DNA, traditional markers can only detect cytoplasmic DNA from a single parent due to differences in template concentration and PCR competition (Wang et al., 2010), resulting in misjudgment

Method used

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  • Method and device for detecting cytoplasmic inheritance
  • Method and device for detecting cytoplasmic inheritance
  • Method and device for detecting cytoplasmic inheritance

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0062] The detection of mitochondrial DNA inheritance in Arabidopsis includes the following steps:

[0063] (1) According to literature reports (Matsushima et al., 2011), Arabidopsis thaliana ecotypes C24 and Col have a SNP site in the genome sequence of the mitochondrial gene matR. Therefore, for this SNP site, Primer Express3.1 software was used to design primers and probes for the male and female materials based on the TaqMan MGB Allelic Discrimination mode.

[0064] Wherein, the obtained mitochondrial genome sequences of C24 and Col are shown in SEQ ID NO: 1 and SEQ ID NO: 2, respectively, and there is a SNP (SNP.C29A) at 29 bp compared between the two.

[0065] The primer sequences before and after the two parents designed for this sequence are identical, as shown in SEQ ID NO: 3 and SEQ ID NO: 4, respectively, the sizes are 26 and 20 bp, respectively, and the size of the amplified fragment is both 68 bp.

[0066] The specific probe sequence designed for C24 is shown in ...

Embodiment 2

[0076] The detection of cucumber mitochondrial DNA genetic mode includes the following steps:

[0077] (1) Comparatively analyze the mitochondrial genome polymorphisms of the parental materials '9930' and 'B', find suitable polymorphic sites, and then design synthetic primers and probes for the polymorphic sites, and the probes span polymorphisms site and label the FAM and HEX fluorophores, respectively. The parental materials 9930 and B are inbred line materials preserved in the laboratory. Their mitochondrial genome sequences were obtained by next-generation sequencing, respectively. The software Bowtie 2 was used to compare with the published reference mitochondrial genome of cucumber 'Calypso' to generate bam files. , and load the bam file into IGV 2.4.5, and compare the effective polymorphism sites according to the sequencing results. Then, using Primer Express 3.1 software, based on the TaqManAllelic Discrimination mode, the primers and probes of the male and female mat...

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Abstract

The invention relates to the field of biotechnology, in particular to a method and equipment for detecting cytoplasmic inheritance. The method for detecting cytoplasmic inheritance provided by the present invention includes: (1) based on the polymorphism of the cytoplasmic nucleic acid sequence of the biological material parental sample, obtaining different polymorphic sites in the biological material parental sample respectively, the Parent samples include paternal samples and maternal samples; (2) based on the different polymorphic sites, use digital PCR technology to obtain the polymorphism results of the cytoplasmic nucleic acid sequences of the offspring samples; (3) based on the offspring As a result of polymorphisms in the cytoplasm of the sample, cytoplasmic inheritance of the biological material is determined. At the same time, a device for detecting cytoplasm inheritance is correspondingly provided. The method and device provided by the invention are used to detect the characteristics of cytoplasm inheritance, with flexible sampling, high resolution, avoiding the competition effect between templates, and extremely high sensitivity.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method and equipment for detecting cytoplasmic inheritance. Background technique [0002] Cytoplasmic inheritance refers to the genetic phenomenon determined by genetic factors outside the nucleus, which is called non-Mendelian inheritance or extranuclear inheritance. Cytoplasmic inheritance in animals is mainly mitochondrial inheritance, while cytoplasmic inheritance in plants includes mitochondrial inheritance and plastid inheritance. In plants, the earliest non-Mendelian inheritance phenomenon was reported in 1909. When Baur et al. studied the inheritance of "leaf spot" in geranium, they found that the inheritance of parental leaf spot in offspring did not show Mendelian inheritance (Baur, 1909). ). Cytoplasmic inheritance patterns are closely related to plant cytoplasmic male sterility, growth retardation caused by plastid or mitochondrial genome mutations, etc. Therefore, i...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6858C12Q1/6895C12M1/34C12M1/00
CPCC12Q1/6858C12Q1/6895C12Q2600/156C12Q2531/113C12Q2563/107
Inventor 沈佳张跃建寿伟松
Owner ZHEJIANG ACADEMY OF AGRICULTURE SCIENCES
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