Prepn process and application of sea island cotton EST SSR marker

A sea-island and marker technology, applied in the preparation of sea-island cotton EST-SSR primer sequences and QTL mapping, can solve the problem of small data volume of African cotton and sea-island cotton

Inactive Publication Date: 2007-08-22
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the amount of data for the cultivated species of African cotton and Sea Island co

Method used

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  • Prepn process and application of sea island cotton EST SSR marker
  • Prepn process and application of sea island cotton EST SSR marker
  • Prepn process and application of sea island cotton EST SSR marker

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] 1, get 36 parts of cotton seed materials (as shown in Figure 2), its genotype is AA13, DD11 AAD12) as the material of genetic diversity analysis, and described material is planted in the field;

[0041] 2. Get the tender green leaves from the field material plant and extract its total DNA. The specific method is with reference to Paterson et al. in 1994 (Paterson A H, Curt L B, Wendel J F, A rapid method for extraction of cotton (Gossypium spp.) genomic DNA suitable for RFLP and PCR analysis. Plant Mol Bil Rep, 1993, 11: 112-127) reported method extraction;

[0042] 3. Utilize the sea island cotton EST-SSR primer sequence among the present invention to analyze, and concrete steps are as follows:

[0043] 3.1 HAU primer marker analysis:

[0044] PCR amplification reaction system is 20μL: 25ng DNA, 4μmol upstream primer, 4μmol downstream primer, 1×buffer (10mM Tris-Hcl, 50mM Kcl, pH8.3), 2mmol Mg2 + , 0.25mmol dNTPs, and 0.8 U Taq polymerase.

[0045] The PCR amplifica...

Embodiment 2

[0058] 1. Parent Emian 22, Pima3-79 (provided by the Cotton Research Institute of the Chinese Academy of Agricultural Sciences) and BC 1 The population is used as the starting material for the construction of the genetic linkage map, and the material is planted in the field;

[0059] 2. Extract its total DNA from the tender green leaves on the field material plant;

[0060] 3. Utilize the sea island cotton EST-SSR primer sequence among the present invention to analyze, and concrete steps are as follows:

[0061] 3.1 HAU primer marker analysis:

[0062] The PCR amplification reaction system was 20 μL: 25ng DNA, 4 μmol upstream primers, 4 μmol downstream primers, 1 × buffer (10mM Tris-Hcl, 50mM Kcl, pH8.3), 2mmol Mg2+, 0.25mmol dNTPs, and 0.8 U Taqpolymerase.

[0063] The PCR amplification reaction program was as follows: denaturation at 94°C for 3 minutes, followed by 34 cycles (denaturation at 94°C for 50 seconds, annealing at 57°C for 45 seconds, extension at 72°C for 1 minut...

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Abstract

The present invention belongs to the field of cotton breeding technology, and is especially the preparing technology of sea island cotton EST-SSR primer sequence and its application in cotton genetic diversity evaluation, molecular genetic linkage construction and TQL location of important cotton characters. The preparation process includes the following steps: establishing cDNA library of sea island cotton strain Pima3-79 fiber development and selecting EST of partial bases; designing SSR lateral wing primer; taking total DNA with plant leaf; amplifying with the designed primer; calculating the polymorphism information content of each pair of primers and performing cluster and main coordinate analysis of taken total DNA; and integrating the amplified EST-SSR to corresponding linkage group and performing TQL location of important cotton characters. The preparation results are also given.

Description

technical field [0001] The invention belongs to the technical field of cotton breeding, and in particular relates to the preparation technology and application of sea-island cotton EST-SSR primer sequences, in particular to the evaluation of cotton genetic diversity, the construction of molecular genetic linkage and the QTL positioning of important cotton traits by using these primer sequences . Background technique [0002] Simple Sequence Repeat (SSR) markers have the advantages of good repeatability, high polymorphism, co-dominant inheritance, abundant quantity, and spread throughout the genome. These advantages make SSR markers one of the most widely used molecular markers. However, according to traditional The method development of SSR markers is not only time-consuming and laborious but also inefficient. [0003] With the development of functional genomics, expressed sequence tags (ESTs) have been sequenced in large quantities because they can be used for the discover...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C40B40/06
Inventor 张献龙林忠旭张艳欣涂礼莉聂以春
Owner HUAZHONG AGRI UNIV
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