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Screening method for hydroxyl carthamin yellow A-containing safflower

A kind of hydroxyl safflower, screening method technology, applied in the field of biological sciences

Inactive Publication Date: 2008-10-01
SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, there is no application of cDNA-AFLP technology combined with BSA method for differential expression analysis of safflower with and without hydroxyl safflower yellow A

Method used

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  • Screening method for hydroxyl carthamin yellow A-containing safflower
  • Screening method for hydroxyl carthamin yellow A-containing safflower
  • Screening method for hydroxyl carthamin yellow A-containing safflower

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Embodiment 1: TDF-8 of the present invention, 9, the separation of 27 fragments

[0038] 1. Determination of F by high performance liquid chromatography, thin layer chromatography and ultraviolet spectrophotometry 2 The content of hydroxysafflor yellow A in the generation population was detected for its presence or absence.

[0039] 2. According to the measurement results, the safflower corollas containing and not containing hydroxysafflower yellow A were taken respectively, the total RNA was extracted, and converted into double-stranded cDNA.

[0040] 3. Construct near-isogenic pools with and without hydroxysafflower yellow A, take 1 μg of cDNA from a single plant containing hydroxysafflower yellow A, a total of 10 strains are mixed to form a pool; another one without hydroxysafflower yellow A A total of 10 cDNA strains were mixed with 1 μg each to form a pool.

[0041] 4. cDNA-AFLP differential expression analysis:

[0042](1) Restriction digestion and linker ligat...

Embodiment 2

[0048] Example 2: Verification of TDF-8, 9, and 27 gene expression by RT-PCR:

[0049] A. Design and synthesize RT-PCR primers according to the sequences of TDF-8 (SEQ ID NO: 1), TDF-9 (SEQ ID NO: 2), and TDF-27 (SEQ ID NO: 3),

[0050] TDF-8 Primer: S8: 5'-GTCCTGAGTAAGCAAGTGTGT-3'

[0051] A8: 5'-ACCATTGATAGCGGTGTCCA-3'

[0052] TDF-9 Primer: S9: 5'-CTCCTTGCGGAACATGAATGCTG-3'

[0053] A9: 5'-AGCGTACCAATTCTGGACCTGAT-3'

[0054] TDF-27 Primer: A27: 5'-AGTAATCTTGGGCATCCACGCTG-3'

[0055] S27: 5'-TTGGAGGATAAAGAGGAGGATCAGG-3'

[0056] B, extract the total RNA of Henan Dahongpao safflower;

[0057] C. Using the safflower total RNA as a template, transcribe it into the first-strand cDNA;

[0058] D, using the first-strand cDNA as a template, adding synthetic primers, and performing PCR amplification;

[0059] E, agarose electrophoresis, ultraviolet color development.

[0060] F. Sequencing.

[0061] The same sequence gene as SEQ ID NO: 1-3 ...

Embodiment 3

[0062] Embodiment 3: the application of this screening method:

[0063] 1. Extract the total RNA of safflower with and without hydroxysafflower yellow A in each producing area, and convert it into double-stranded cDNA.

[0064] 2. Using the combination of E10M8 and E9M3 primers, the cDNA-AFLP differential expression analysis was performed on safflowers containing hydroxy safflower yellow A and those without safflower yellow A by the method of implementation 1. Flower, Changji safflower, Xinxiang safflower, Heze safflower, Bozhou safflower, Jianyang safflower, Kaifeng safflower, Zhengzhou safflower, Chengdu safflower, Weishan safflower, Jiuquan safflower, Kunming safflower get 3 unique However, Ruoqiang safflower, Tacheng safflower, Urumqi safflower, Qingxu safflower and Hainan safflower were not obtained.

[0065] 3. The above-mentioned safflower varieties containing 3 specific fragments all contain hydroxysafflower yellow A as determined by HPLC method; while the varieties w...

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Abstract

The invention relates to the field of bioscience technology; as main active ingredients of safflower, carthamin yellow A with hydroxyl has the content which directly influences on the quality of the safflower. The invention takes pure-line parent of the carthamin yellow A with hydroxyl and without hydroxyl and F<1>, F<2> generations generated by mating as the materials; in accordance with the presence of carthamin yellow A with hydroxyl in each plant, carthamin yellow A with hydroxyl presence two ponds are established by applying BAS method and cDNA-AFLP method is used to analyze the expression properties of the both; differential expression genes in bound up with the carthamin yellow A with hydroxyl are separated; correctness of property gene type and cDNA-AFLP method is verified via RT-PCR. The specificity DNA segment obtained by the invention includes the nucleotide sequence shown as SEQ ID NO: 1-3. The invention applies the specificity DNA segment to screening the safflower breed of the carthamin yellow A with hydroxyl in order to realize to provide good conditions for oriented adjustment and control of the high quality properties of safflower.

Description

technical field [0001] The invention relates to the technical field of biological sciences, in particular to a screening method for safflower containing hydroxyl safflower yellow A Background technique [0002] Carthmus tinctorius L. is the dry tubular flower of Carthmus tinctorius L., which is widely used clinically. The main chemical components are flavonoids, indoles, lignans, alkyl glycols and so on. Hydroxysafflor yellow A (HSYA for short) in flavonoids is the main active ingredient of safflower, which has various pharmacological effects such as anti-myocardial ischemia, inhibition of platelet aggregation, and anti-oxidation. Concern (Shi Feng, Liu Yanwen, Research Progress on Chemical Constituents and Pharmacology of Safflower, Shi Zhen Guo Yi Guo Yao. 2006, 17(9): 1666-1667). Therefore, the content of hydroxysafflower yellow A directly affects the quality of safflower. The study of functional genes related to safflower quality is of great significance to improve th...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 郭美丽冯娜唐洁
Owner SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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