Pinewood nematoda detection reagent-box and detection method

• Main Authors: BENYUAN YU, MAOSONG LIN, JINCHENG WANG
• Format: Patent
• Language: Chinese; English
• Published: 15.09.2004
• Edition: 7
• Subjects: BEER; BIOCHEMISTRY; CHEMISTRY; COMPOSITIONS OR TEST PAPERS THEREFOR; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL ORENZYMOLOGICAL PROCESSES; ENZYMOLOGY; INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIRCHEMICAL OR PHYSICAL PROPERTIES; MEASURING; MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEICACIDS OR MICROORGANISMS; METALLURGY; MICROBIOLOGY; MUTATION OR GENETIC ENGINEERING; PHYSICS; PROCESSES OF PREPARING SUCH COMPOSITIONS; SPIRITS; TESTING; VINEGAR; WINE

Two pairs of primer are designed: Pl/P3 is an idiocratic primer for pine wood nematode; P2/P3 an idiocratic primer for quasi pine wood nematode. The kit includes 1.25ml solution I,1ml solution II, 100U tag enzyme, and 50ul for each contrapositive DNA of wireworm. The solution I is as following constitutes: Worm Lysis Buffer, 500mM KCl, 100mMTris-ClpH 8.0, 15mM MgCl2, 10mMDTT, 4.5% Tween20, 0.1%gelatin, 0.2ug/ul protease K. The solution II is as following constitutes: 2xPCR reaction Buffer, 4.0mM MgCl2, 0.2mM dNTP, 0.8 mu MP1, P2, P3. The invented kit possesses high specificity, sensitiveness and stability, providing fast, sensitive and specific technical method. 本发明松材线虫检测试剂盒及其检测方法属于农林作物防病治病及植物检疫范畴。设计了两个引物对，P1/P3为松材线虫特异性引物对，P2/P3为拟松材线虫特异性引物对。试剂盒：1.25ml溶液I，(Worm Lysis Buffer，500mM KCl，100mM Tris-Cl pH 8.0，15mM MgCl#-[2]，10mM DTT，4.5％Tween 20，0.1％gelatin，0.2ug/ul蛋白酶K)，1ml溶液II(2×PCR反应Buffer，4.0mM MgCl#-[2]，0.2mMdNTP，0.8μM P#-[1]、P#-[2]、P#-[3])，100U taq酶，线虫对照DNA各50ul。该检测试剂盒具较强的特异性、灵敏性及稳定性，提供了快速、灵敏、特异的技术方法。