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Method for analyzing differential expression of cotyledon senescence-associated gene of brown cotton by use of cDNA-SRAP (complementary deoxyribonucleic acid-sequence-related amplified polymorphism) technique

A technology of technical analysis and genetic difference, applied in the fields of biochemical equipment and methods, microbial determination/inspection, etc., to achieve the effect of low cost, high efficiency and deepening of understanding

Inactive Publication Date: 2012-07-25
ANHUI AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Ethylene has now been identified as a potent senescence promoter, but its effect on brown cotton cotyledon senescence has rarely been reported

Method used

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  • Method for analyzing differential expression of cotyledon senescence-associated gene of brown cotton by use of cDNA-SRAP (complementary deoxyribonucleic acid-sequence-related amplified polymorphism) technique
  • Method for analyzing differential expression of cotyledon senescence-associated gene of brown cotton by use of cDNA-SRAP (complementary deoxyribonucleic acid-sequence-related amplified polymorphism) technique
  • Method for analyzing differential expression of cotyledon senescence-associated gene of brown cotton by use of cDNA-SRAP (complementary deoxyribonucleic acid-sequence-related amplified polymorphism) technique

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Embodiment 1 ethephon induces brown cotton cotyledon senescence

[0037] 1 material

[0038] Using the brown cotton "Brilliant Choice No. 1" that had germinated for 20 days as the material, the leaves were sprayed with ethephon at a concentration of 0, 10, 50, 150, 250, 500, and 1000 ppm, and the cotyledons were taken after 24 hours of treatment. Take 6 plants and repeat three times. Store in a -70°C refrigerator after taking pictures.

[0039] 2 methods

[0040] For enzyme extraction and activity determination, refer to "Principles and Techniques of Plant Physiological and Biochemical Experiments" edited by Wang Xuekui (2006). The POD activity was measured by the guaiacol method, the SOD activity was measured by the nitro blue tetrazolium (NBT) method, and the CAT activity was measured by ultraviolet spectrophotometry. The specific measurement method is as follows:

[0041] Determination of peroxidase (POD) activity

[0042] Determination of peroxidase activity b...

Embodiment 2

[0070] Example 2 Ethephon Treatment Effects on Gene Expression

[0071] Step 1, first use concentrated sulfuric acid with a mass fraction of 98% to delint and disinfect the brown cottonseed of "Zongcaixuan No. 1", and plant it at 10cm 3 In the plastic culture pot, the cultivation was carried out in the light incubator, and the light intensity was 45μmol*m -2 *s -1 , photoperiod (16 / 8h), and then cultivated in a light incubator at 27°C ± 2°C at a constant temperature. The brown cotton "Wonderful Choice No. 1" that germinated for 20-25 days was used as the material, and the leaves were sprayed with ethephon at a concentration of 500ppm. After 18-24 hours of treatment, the cotyledons were taken, treated with liquid nitrogen, and stored in a -70°C refrigerator for later use;

[0072] Step 2, extracting total RNA from cotton cotyledons. The extraction of total RNA adopts the improved CTAB method, and the extracted total RNA is detected by ultraviolet spectrophotometer, and the c...

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Abstract

The invention discloses a method for analyzing differential expression of cotyledon senescence-associated genes of brown cotton by use of a cDNA-SRAP (complementary deoxyribonucleic acid-sequence-related amplified polymorphism) technique. The method comprises the following steps: firstly, spraying ethylene with concentration of 450-550ppm on a to-be-analyzed material to process for 18-24h and extracting total RNA (ribonucleic acid) in cotyledon; and performing cDNA-SRAP PCR (polymerase chain reaction), recovering and purifying specific bands obtained in a reaction product and analyzing functions of sequences. According to the invention, cDNA-SRAP technique is utilized to analyze differential expression of the related genes on a molecular level, so as to explore the molecular mechanism of cotyledon senescence of brown cotton, deepen understanding of the leaf senescence of brown cotton, and provide a theoretical basis for artificial control of leaf senescence process of colorful cotton.

Description

technical field [0001] The invention belongs to the field of biotechnology; more specifically, the invention relates to the analysis of the influence of ethephon treatment on brown cotton cotyledon senescence and the differential expression analysis of related genes by using cDNA-SRAP technology. Background technique [0002] In cotton production, premature leaf senescence is one of the important factors leading to cotton yield reduction. Premature senescence of leaves in cotton fields can lead to yield loss of about 10%, and serious cases can reach more than 20%. The premature aging of cotton leaves not only affects the yield of cotton, but also leads to the shortening of cotton fiber length, the reduction of fiber strength, and the decrease of fiber uniformity, thereby reducing the yarn quality of cotton fiber. Natural colored cotton, because its fibers have natural brown, green and other colors, does not need artificial dyeing in the textile processing process, and avoid...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 郭宁王旺华林毅樊洪泓高俊山姜家生
Owner ANHUI AGRICULTURAL UNIVERSITY
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