Details
Store in the dark at -20℃, transport at 2-8℃, valid for one year. Avoid repeated freezing and thawing.
Kit Features:
1. Specific detection of Mycoplasma bovis without cross-reaction with other organisms.
2. High sensitivity, the lowest detection limit is equivalent to about 1 CFU in the reaction solution.
3. The DNA polymerase used has the characteristics of strong anti-inhibition ability and good thermal stability; using hot start method, it can Inhibit non-specific amplification and reduce background fluorescence.
4, with positive control sample, which can be used to test the effectiveness of the kit.
5. With UDG enzyme and dUTP, it can reduce the contamination of residual DNA.
Mycoplasma bovis is the causative agent of bovine infectious mycoplasma pneumonia . It is the most important pathogenic bovine parasitic mycoplasma. It has a global epidemic and can cause pneumonia, arthritis, mastitis, Otitis media, conjunctivitis, reproductive disorders, miscarriage and other diseases have an incidence rate of 60%-100%. The mortality rate is not high, but sometimes it causes great economic losses. Mycoplasma bovis can be found in both healthy and diseased cattle. Its disease manifestations and response to treatments and vaccines often vary, making diagnosis and control difficult, so its prevalence is often underestimated.
The detection methods of Mycoplasma bovis include: in vitro culture method, serological method, and Genetic methods. The in vitro culture conditions of Mycoplasma bovis are relatively harsh and take a long time, sometimes even two weeks, and require specific methods for identification, which are prone to interference from other respiratory pathogens. Serological methods can only detect the infection after a period of time.The detection time is delayed. In addition, there are antigenic and genetic variations between laboratory strains and strains isolated in the field, which interferes with the detection. Therefore, the highly sensitive and specific PCR method is a better choice because its detection time is short and the results can be obtained in just a few hours. Compared with the ordinary PCR method, the quantitative PCR method can not only accurately quantify, but also be more convenient to operate and less affected by environmental pollution.
Due to interference from Mycoplasma agalactiae and other closely related mycoplasmas, it is based on 16S rRNA Genetic PCR methods often cannot accurately identify M. bovis. The PCR method based on the DNA replication repair gene uvrC can specifically distinguish these mycoplasmas, and the gene is very stable and rarely mutates. Therefore, this kit selects the uvrC gene as the target for PCR identification of Mycoplasma bovis. After verification by BLAST, It has very good specificity and no cross-reactivity with the genomes of other organisms. By detecting 10 closely related mycoplasma species, 21 common bacteria, and 19 healthy bovine lung tissues, no false positive results were found, indicating that this kit is specific for Mycoplasma bovis. Testing of 385 tissue samples showed that the test results of this kit were completely consistent with those of the culture method.
This kit contains hot-start DNA polymerase, dNTP, primers, probes, positive controls, ROX dye, and UDG to prevent residual DNA contamination. Users only need to prepare DNA samples and use them.
The DNA polymerase used in this kit is derived from extremothermophilic bacteria, which has been modified to have stronger resistance to inhibition and thermal stability than ordinary Taq enzymes. By binding specific monoclonal antibodies, DNA polymerase activity is inhibited at room temperature. During the thermal denaturation phase of the PCR cycle, the antibodies are heated and removed, and the DNA polymerase activity is restored at this time, thus obtaining the "hot start" function. Hot start can reduce the contamination of residual DNA and improve PCR amplification efficiency, sensitivity and target fragment yield.
-20℃避光保存，2-8℃运输，有效期一年。避免反复冻融。
试剂盒特点：
1， 特异性检测牛支原体，与其他生物无交叉反应。
2， 灵敏度高，最低检测极限相当于反应液中约含1个CFU。
3， 使用的DNA聚合酶具有抗抑制能力强和热稳定性好的特点；采用热启动方式，可抑制非特异性扩增，降低背景荧光。
4， 带有阳性对照样品（组分C），可用于检验试剂盒有效性。
5， 带有UDG酶和dUTP，可降低残留DNA的污染。
牛支原体（Mycoplasma Bovis）是牛传染性支原体肺炎（又称牛烂肺病）的病原体，是最重要的致病性牛寄生支原体，呈全球流行趋势，可引起肺炎、关节炎、乳腺炎、中耳炎、结膜炎、生殖紊乱和流产等多种疾病，发病率达到60%-100%，病死率不高，但有时会造成很大的经济损失。在健康或患病的牛身上均可发现牛支原体，其疾病表现和对治疗和疫苗的反应常发生变化，诊断和控制较为困难，因此其流行程度常被低估。
牛支原体的检测方法包括：体外培养法，血清学方法，和遗传学方法。牛支原体的体外培养条件较为苛刻，用时较长，有时甚至要两周时间，并且需要特定的方法来进行鉴定，容易受到其他呼吸道病原体的干扰。血清学方法则必须在感染一段时间之后才能检测，存在检测时间的延迟，此外，实验室菌株和野外分离获得的菌株存在抗原和遗传学上的变异，对检测形成了干扰。因此，高灵敏度和高特异性的PCR法是更好的选择，其检测时间短，仅需几个小时即可获得结果。定量PCR法与普通PCR法相比，不仅可以精确定量，而且操作更为方便，更少受环境污染的影响。
由于受到无乳支原体及其他一些近亲支原体的干扰，基于16S rRNA基因的PCR方法往往无法准确鉴别牛支原体。基于DNA复制修复基因uvrC的PCR方法可以对这些支原体进行特异性区分，而且该基因十分稳定，很少发生变异，因此本试剂盒选择uvrC基因作为靶点对牛支原体进行PCR鉴定，经BLAST验证，具有非常好的特异性，与其他生物的基因组无交叉反应。通过检测10种近亲支原体和21种常见细菌，以及19个健康牛肺组织，未见假阳性结果，可见本试剂盒对牛支原体具有特异性。对385个组织样品的检测表明，本试剂盒与培养法检测结果完全一致。
本试剂盒包含热启动DNA聚合酶、dNTP（以dUTP代替dTTP）、引物、探针、阳性对照品、ROX染料，和用以防止残余DNA污染的UDG（Uracil- DNA Glycosylase，尿嘧啶-DNA-糖基酶），用户只需准备好DNA样品即可使用。
本试剂盒采用的DNA聚合酶源自极端嗜热菌（Thermus thermophilus），经改造后较普通的Taq酶具有更强的抗抑制能力和热稳定性。通过结合特异性单克隆抗体，在室温下DNA聚合酶活性被抑制。在PCR循环的热变性阶段，抗体受热被去除，此时DNA聚合酶活性恢复，因此获得“热启动”功能。热启动可减少残余DNA的污染，提高PCR扩增效率、灵敏度和目的片段产量。
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