Lophopyrum elongatum genome-specific molecular markers and application thereof

A technology of R. longarum and molecular marker, which can be applied in the field of crop genetics and breeding, and can solve the problems such as the lack of research on R. longarum.

Inactive Publication Date: 2012-09-19
YANGZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] 2. Wheat head blight resistance
At present, ISSR markers have been widely used in the identification of various plant varieties, genetic mapping, gene mapping, genetic diversity, etc., but have not been studied in Echinopsis elongatum.

Method used

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  • Lophopyrum elongatum genome-specific molecular markers and application thereof
  • Lophopyrum elongatum genome-specific molecular markers and application thereof
  • Lophopyrum elongatum genome-specific molecular markers and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0154] Embodiment 1 experimental material and PCR primer sequence

[0155] (1) Experimental materials

[0156] Common wheat China Spring (CS), diploid Enopyrum elongatum (EE), China Spring-Etnopyrum elongatum disomic addition lines: DA1E, DA2E, DA3E, DA4E, DA5E, DA6E, DA7E (The above materials are provided by Canadian Agriculture Donated by Dr.Fedax from the Ministry of Science and Technology, the original document of the material is "Disomic and ditelosomic additions of diploid Agropyron elongatum chromosomes to Triticum aestivum"); Yangmai 158 (Y158, used by the Agricultural Science Institute of Lixiahe District, Jiangsu Province Yangmai No. 4 / ST1472 / 506 bred, the variety registration number is GS02001-1997), Yangmai 16 (Y16, that is, Yang 0-126 is a hybrid of Yang 91F138 and Yang 90-30 used by the Agricultural Science Institute in Lixiahe, Jiangsu), Ningmai 13 (N13, Namely Ning 0078, National Examination Mai 2006004, selected from the Ningmai No. 9 line by the Institute o...

Embodiment 2

[0160] The extraction of embodiment 2 genome DNA

[0161] The test material was grown to two leaves and one heart stage, and the genomic DNA was extracted by SDS method. The steps are as follows:

[0162] (1) Take the young leaves (about 0.1g), cut them into pieces and put them into a 2ml centrifuge tube, cool them in liquid nitrogen, and grind them to powder with a grinding rod;

[0163] (2) Place the centrifuge tube at room temperature to cool slightly, add 700 μl of buffer A, mix gently, then bathe in water at 65°C for 20 minutes, and mix by inverting up and down every 5 minutes;

[0164] Buffer A: NaCl 29.2g

[0165] 1M Tris-HCl 100ml

[0166] EDTA 18.6g

[0167] SDS 15g

[0168] Dilute the volume to 1L with ddH2O and use it after sterilization.

[0169] (3) Take it out and cool it down to room temperature, add 350 μl of phenol and chloroform each, turn it upside down, mix well, and extract for 5 minutes;

[0170] (4) 12000rpm, centrifuge ...

Embodiment 3

[0177] Embodiment 3 suppresses the construction of subtractive hybridization library

[0178] Use the suppression subtractive hybridization (SSH) technique to construct a hybrid library, the specific steps are as follows:

[0179] (1) DNA digestion and ligation and efficiency detection. The DNA of Echinopsis elongatum and Chinese spring were digested with Rsa I enzyme (Clontech Company), the total volume of the enzyme digestion reaction was 50 μL, incubated at 37°C for 24 h, and 1.5 μL of Rsa I was added once in the middle. The digested product of E. elongatum was ligated with linker 1 and linker 2R, respectively. After the ligation reaction, check the ligation efficiency of the ligated product, and carry out 2 rounds of hybridization and 2 rounds of PCR amplification for the ligated product, and the related operations are as follows: PCR-Select TM cDNA Subtraction Kit instructions were performed. The electrophoresis results of the RsaⅠ digested genomic DNA of Echinopsis e...

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Abstract

The invention belongs to the field of crop genetic breeding, in particular 36 Lophopyrum elongatum genome-specific molecular markers. The 36 genome-specific molecular markers are obtained by the following steps: according to a nucleotide sequence of 18srRNA gene disclosed by National Center of Biotechnology Information (NCBI), designing primers in the sequence between two RsaI loci, removing homologous sequences between two genome DNA by using a suppression subtractive hybridization technology, enriching differential sequences, obtaining Lophopyrum elongatum specific DNA segments, sequencing the DNA segments, and redesigning primers according to sequences without homology with wheat. The markers can be used for identifying translocation lines in the process of transferring chromosome segments from Lophopyrum elongatum to wheat, can be used for linkage analysis of scab-resistant gene, and can serve as specific molecular markers for identifying chromatin of the Lophopyrum elongatum to be applied to scab-resistant and stress-resistant breeding of wheat.

Description

technical field [0001] The invention belongs to the field of crop genetics and breeding, and in particular relates to genome-specific molecular markers of Echinopsis elongatum and applications thereof. Background technique [0002] 1. Wheat breeding goals and its wild relatives [0003] Wheat is the food crop with the largest planting area in the world and its total output is second only to corn. In the past half century, the world's total wheat production has more than tripled, and excellent wheat varieties have played a decisive role in increasing wheat production. At present, wheat breeding research mainly focuses on four aspects: increasing wheat yield and reducing production cost; molecular marker-assisted breeding; wheat stress resistance breeding; wheat nutritional quality (He Zhonghu et al., 2006). In order to accomplish the above breeding goals, the genetic resources of wheat itself are obviously insufficient, but there are a large number of favorable genetic reso...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/11C12Q1/68
Inventor 陈建民陈士强葛江燕高勇
Owner YANGZHOU UNIV
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