Sugarcane genome endogenous reference gene fructose-6-phosphate 2-kinase gene PCR (polymerase chain reaction) primer sequences and amplification method

An internal standard gene and primer sequence technology, applied in recombinant DNA technology, DNA/RNA fragments, DNA preparation, etc., can solve the problem of no research report on copy number, and achieve the effects of easy determination, strong specificity, and wide applicability

Inactive Publication Date: 2015-02-04
FUJIAN AGRI & FORESTRY UNIV
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  • Description
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  • Application Information

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Problems solved by technology

There are also many studies on the fructose-6-phosphate 2-kinase gene that produces this enzyme, but there is no research report on the copy number of this gene in the genome of sugarcane species

Method used

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  • Sugarcane genome endogenous reference gene fructose-6-phosphate 2-kinase gene PCR (polymerase chain reaction) primer sequences and amplification method
  • Sugarcane genome endogenous reference gene fructose-6-phosphate 2-kinase gene PCR (polymerase chain reaction) primer sequences and amplification method
  • Sugarcane genome endogenous reference gene fructose-6-phosphate 2-kinase gene PCR (polymerase chain reaction) primer sequences and amplification method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] (1) PCR amplification of sugarcane fructose-6-phosphate 2-kinase gene

[0022] Fructose-6-phosphate 2-kinase gene PCR primer sequence is: ScF2KP -F:CCTGGGAGGATTGTCTTCTTC; ScF2KP -R: ATCACCACCGATTCTTCCTCT.

[0023] PCR reaction system: 1.0 U Taq DNA polymerase, 1×PCR buffer solution, 0.25 mM dNTP, 3.0 mM MgCl 2 , primer ScF2KP -F and ScF2KP Each of -R was 0.25 mM, DNA template was 25 ng, and the total volume of the reaction system was 25 μl.

[0024] PCR amplification conditions: pre-denaturation at 95°C for 6 min; denaturation at 94°C for 30 s; annealing at 56°C for 30 s, extension at 72°C for 35 s, a total of 35 cycles, and a final extension at 72°C for 7 min.

[0025] (2) Electrophoresis detection of PCR product of sugarcane fructose-6-phosphate 2-kinase gene and verification of product specificity and universality

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Abstract

The invention discloses sugarcane genome endogenous reference gene fructose-6-phosphate 2-kinase gene PCR (polymerase chain reaction) primer sequences and an amplification method. The primer sequences are composed of ScF2KP-F and ScF2KP-R. The PCR amplification method comprises a reaction system and amplification conditions. The invention provides a genome endogenous reference gene for PCR detection and species specific identification of transgenic sugarcane, and meanwhile, provides insurance for detection precision and result accuracy of the transgenic sugarcane.

Description

technical field [0001] The invention belongs to the field of biological technology, and in particular relates to a PCR primer sequence and an amplification method of an internal standard gene fructose-6-phosphate 2-kinase gene in sugarcane genome. Background technique [0002] Transgenic technology is becoming more and more mature, and the types of genetically modified sugarcane are becoming more and more diverse. At present, herbicide-resistant, insect-resistant, and disease-resistant genetically modified sugarcane has been field tested in 14 countries including the United States, Australia, India, and Brazil, creating conditions for the commercial cultivation of genetically modified sugarcane. In my country, sugarcane gene cloning and transgenic research only started in the mid-1990s. Although it started late, it has made rapid progress. The transgenic sugarcane that has been obtained includes insect-resistant genetically modified sugarcane, drought-resistant genetically m...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/11C12N15/10
Inventor 陈平华林冰陈忠伟陈利平施桂姣张卓项慰刘迪夏峰王恒波高三基许莉萍陈如凯
Owner FUJIAN AGRI & FORESTRY UNIV
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