摘要：本研究前期发现，生防假单胞菌（Pseudomonas protegens）Pf-5菌株对根结线虫具有致死活性。为了研究该生防菌对线虫的致死机理，本研究以模式线虫秀丽小杆线虫(Caenorhabditis elegans)为作用对象，开展相关研究工作。研究表明，Pf-5菌株对秀丽小杆线虫具有很强的致死活性。利用荧光蛋白GFP标记的Pf-5(gfp)菌株的观察结果表明，Pf-5菌株能定殖在线虫的口针和食道部位。我们进一步利用pK18mobsacB无标记载体，构建了Pf-5△fitD突变体。结果表明，Pf-5△fitD虽然也能定殖在线虫的口针和食道部位，但是杀线虫活性几乎完全丧失。不同浓度的菌体处理结果表明，Pf-5菌体浓度在107CFU/mL及以上时，都具有杀线虫活性，而Pf-5△fitD在相应处理浓度下，杀线虫活性几乎完全丧失。当菌体浓度在108CFU/mL时，Pf-5菌株处理6 h，线虫的死亡率接近100%，当处理时间为24 h时，线虫完全死亡。而Pf-5△fitD突变体在108CFU/mL时，各个时间段线虫的死亡率都只有10%左右。该研究表明，Pf-5菌株的杀线虫活性与fitD基因有关，为进一步的机理研究提供了研究材料。82391

毕业论文关键词：生防假单胞菌；杀线虫活性；机理研究

Functional identification the nematocide substance produced by Pseudomonas protegens

Abstract ：Previous studies have　showed　that Pseudomonas protegens Pf-5 strain has lethal activity against root-knot nematodes。 In order to study the biocontrol mechanism of Pf-5, the model nematodes Caenorhabditis elegans was used in this study。 The results had showed that the Pf-5 strain has a strong lethal activity against C。 elegans。 The results also indicated that Pf-5 strain labeled with the fluorescent protein GFP could colonize in the mouth and the esophagus of the nematodes。 We used the pK18mobsacB unlabeled vector to construct the Pf-5△fitD mutant。 The results showed that Pf-5△fitD mutant was also able to colonize the mouth and esophagus of the nematode, but the nematode activity was almost completely lost。 The results of different concentrations of bacteria showed that the Pf-5 strain had a nematocidal activity at 107CFU/mL, but Pf-5△fitD was almost completely lost this activity at the corresponding treatment concentration。 When nematodes were treated with 108CFU / mL of Pf-5 for 6 h, the mortality of nematode was close to 100%。 When the time of treatment was prolonged to 24 h, the nematodes died completely。 While the mortality of the Pf-5△fitD mutant was only about 10% at 108CFU/mL。 This study shows that the nematocidal activity of the Pf-5 strain is related to the fitD gene and provides a study material for further mechanism studies。 

Key words: Pseudomonas protegens；The activity of killing nematode；Mechanism 

目  录

摘要 1

关键词 1

Abstract 1

Key words 1

引言 1

1材料与方法 2

1。1供试菌株与质粒 2

1。1。1供试菌株与质粒 2

1。1。2菌株的培养与保存 3

1。2培养基与抗生素 3

1。2。1培养基 3

1。2。2抗生素 3

1。3试验试剂与器材 3

1。3。1试验试剂 3

1。3。2试验器材 4

1。4供试线虫 4

2试验方法