Improved mitochondrial genome complete sequence determination method

A mitochondrial genome and measurement method technology, applied in the direction of microbial measurement/inspection, biochemical equipment and methods, etc., can solve problems such as interference, achieve the effect of ensuring the success of PCR amplification, reducing costs and technical difficulties

Inactive Publication Date: 2010-11-03
HAINAN UNIVERSITY
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Problems solved by technology

But this approach is susceptible to interference from mitochondrial pseudogenes

Method used

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  • Improved mitochondrial genome complete sequence determination method
  • Improved mitochondrial genome complete sequence determination method
  • Improved mitochondrial genome complete sequence determination method

Examples

Experimental program
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Effect test

Embodiment 1

[0044] Example 1: Determination of the Complete Sequence of the Mitochondrial Genome

[0045] 1. From the query and screening on the NCBI website, the complete mitochondrial genome sequence of the wrasse family sea pig was obtained, and the sequence was used as a template to design PCR primers for the determination of the complete mitochondrial genome sequence of the wrasse

[0046] 2. Using the software Primer Premier 5.0 to design PCR primers, the results are shown in Tables 1 and 2 below:

[0047] Table 1 The first batch of PCR primers and the second pair of mosaic primers added due to poor amplification effect

[0048]

[0049] Table 2 The second batch of PCR primers for the complete sequence determination of the mitochondrial genome

[0050]

[0051] All designed primers were synthesized by Shanghai Sangon Biotechnology Co., Ltd.

[0052] 3. Total DNA extraction and PCR amplification:

[0053] 1) Total DNA extraction was slightly modified according to the method ...

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Abstract

The invention discloses an improved mitochondrial genome complete sequence determination method based on the conventional PCR technology, which aims at providing a low-cost, fast and new mitochondrial genome complete sequence determination method having a simple experimental technology. The method is characterized by the screening of templates and the design of PCR primers, i.e determining the minimal homology of mitochondrion complete sequences of closely-related species which can be used as the templates; carrying out primer design in two batches, and introducing a nested-PCR method to the sequence amplification process; and presenting general rules for the design of the PCR primers. The method is mainly used for mitochondrial genome complete sequence determination, and serves basic scientific researches, such as genetics, molecular biology and the like.

Description

technical field [0001] The invention relates to an improved method for determining the complete sequence of the mitochondrial genome and its application in determining the complete sequence of the mitochondrial genome. Background technique [0002] Mitochondria is a basic and important organelle that exists in most eukaryotic cells. It is the place where cells undergo oxidative phosphorylation. People attribute the genetic information system of mitochondrial genome (mtDNA) to true The second genetic information system of nuclear cells, or extranuclear genes and their expression systems. The complete sequencing of mitochondrial genomes of a large number of different animals shows that animal mitochondrial genes are composed of double-stranded circular DNA molecules of approximately 15-20kb. Mitochondrial genes encode approximately 22 tRNAs, 2 large and small rRNAs and 13 hydrophobic protein polypeptides, which include subunits of the enzyme complex associated with the inner m...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
Inventor 尹绍武齐兴柱陈国华张本骆剑霍蕊
Owner HAINAN UNIVERSITY
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