Rape transgenosis detecting kit

A detection kit and transgenic technology, which are applied in the determination/inspection of microorganisms, biochemical equipment and methods, etc., can solve the problem of time-consuming, labor-intensive and high cost

Active Publication Date: 2014-04-30
SICHUAN HUAHAN TRIO BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] The purpose of the present invention is to overcome the time-consuming, labor-intensive and high-cost defects of the prior art, and provide

Method used

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  • Rape transgenosis detecting kit
  • Rape transgenosis detecting kit
  • Rape transgenosis detecting kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0075] The rapeseed transgenic detection kit in this example consists of a primer combination and a membrane chip. The primer combination is a combination of 11 DPO primers (Dual Priming Oligonucleotide, dual priming oligonucleotide primers). The base sequence of each pair of primers is 5'-3' As follows, wherein base I is inosine nucleotide:

[0076] EPSPS: Forward primer: CTTTCTGGAACCGTCCGTATIIIIIGTGACAA

[0077] Reverse primer: CCAAGTATCACCTTCCTTACIIIIIICTGGCAC;

[0078] GOX: Forward primer: CTGCTGCTCCTAACTGGAAGIIIIIITCACGTT

[0079] Reverse primer: GGTCATTGGAGCACCAGTCAIIIIIIAGGTGAC;

[0080] PAT: Forward primer: TGAGACGTCTACAGTGAACTIIIIIACAGAGC

[0081] Reverse primer: CTTCCAGGGCCCAGCGTAAGIIIIICCAGCCA;

[0082] NPTII: Forward primer: CGAAGTGCCGGGGCAGGATCIIIIITCATCTC

[0083] Reverse primer: CGCGAGCCCCTGATGCTCTTIIIIIAGATCAT;

[0084] BAR: forward primer: GACGACCTCGTCCGTCTGCGIIIIIGCTATCC

[0085] Reverse primer: CAGCAGGTGGGTGTAGAGCGIIIIICCCAGTC;

[0086] BARNASE: Forw...

Embodiment 2

[0154] The rapeseed transgenic detection kit in this example consists of a primer set, a membrane chip and auxiliary materials. The auxiliary materials are deoxyribonucleoside triphosphate (dNTP), EX-Taq polymerase (Polymerase) and a positive oligonucleotide with a biotin mark at the 5' end. Nucleic acid single-stranded DNA, positive oligonucleotide with biotin mark at the 5' end The base sequence of the single-stranded DNA is biotin-5'- GCTTGTACGTAGT

[0155] GATTCTCCCTGACTGCCTCTAACTGTTCGAATACTCATCCTTGCTA -3'.

[0156] The extracted rapeseed genomic DNA is the transgenic rapeseed line HCN92, and the detection results are as follows: figure 2 Shown in B.

[0157] All the other are the same as embodiment one.

Embodiment 3

[0159] The rapeseed transgenic detection kit in this example consists of a primer set, a membrane chip, excipients and liquid preparation, the liquid preparation is 10×PCR buffer, pre-hybridization liquid, hybridization liquid, washing liquid, blocking liquid, streptavidin-labeled spicy Root peroxidase and tetramethylbenzidine (TMB) chromogenic solution, among which, the prehybridization solution is 5×SSC, 0.1% SDS and 10×Denhardt’s; the hybridization solution is 5×SSC, 0.1% SDS, 5×Denhardt’s , 50% deionized formamide and 100ug / ml yeast tRNA; washing solutions were wash solution 1: 2×SSC and 0.1% SDS, wash solution 2: 0.5×SSC and 0.1% SDS, wash solution 3: 100 mM Tris- HCl, pH 7.5, 150 mM NaCl, Wash 4: 100 mM Tris-HCl, pH 9.5, 100 mM NaCl and 100 mM MgCl 2 ; Blocking solution is 3% BSA, 100 mM Tris-HCl, pH7.5, 150 mM NaCl; tetramethylbenzidine chromogenic solution is tetramethylbenzidine chromogenic solution A solution: 200mM sodium citrate, pH5. 4. 0.2mg / ml Tetramethylbenzid...

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Abstract

The invention relates to a rape transgenosis detecting kit in the field of detecting articles. The rapeseed transgenosis detecting kit comprises a primer combination and a membrane chip, and is characterized in that the primer combination is a 11-double-starting oligonucleotide primer combination, the membrane chip refers to 11 groups of probe sequences and one group of positive control probe sequences, which are fixed on the surface of a supporting film, and the 5' end of each probe is provided with an amino marker; asymmetric PCR (polymerase chain reaction) amplification primers, which are Tag primers with biotin marks at 5'ends, are included in the primer combination. The rapeseed transgenosis detecting kit provided by the invention overcomes the defects of time and energy waste and high cost in the prior art, the rape transgenosis detecting kit provided is simple to operate, is quick and sensitive, is high in detection flux, has visual detection results and is low in cost.

Description

Technical field [0001] The invention relates to the field of detection supplies, in particular to a rapeseed transgene detection kit. Background technique [0002] Rapeseed is a Brassica plant in the family Brassicaceae, and it is the earliest and most widely studied crop in the study of transgenic crops. In 1985, Ooms et al. used the Agrobacterium-mediated method to successfully cultivate the first transgenic Brassica napus. Since then, major multinational companies have bred a variety of transgenic rapeseed strains, and after 1995, they have passed biosafety assessments in North America, Japan and other places and approved commercial planting. As of 2010, the planting area of ​​transgenic rapeseed in the world has exceeded 7 million hectares 2 , accounting for 23% of the total planting area of ​​rapeseed in the world, ranking fourth in the world after genetically modified soybeans, corn, and cotton. Rapeseed is an important oil crop, and more than half of my country's c...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
CPCC12Q1/6837C12Q1/686C12Q2537/143C12Q2531/107C12Q2565/501
Inventor 黄广平秦子苏王勇强
Owner SICHUAN HUAHAN TRIO BIOTECH CO LTD
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