阅读说明：本技术 一种培养基及其用于检测大肠杆菌的应用 (Culture medium and application thereof in detection of escherichia coli ) 是由 刘玲 董铭 董绍俊 于 2019-12-23 设计创作，主要内容包括：本发明涉及一种培养基及其用于检测大肠杆菌的应用,属于大肠杆菌检测技术领域。本发明的培养基包括酶底物1为4-甲基伞形酮-β-D-葡萄糖醛酸苷、营养物2为胰蛋白胨或蛋白胨和贮存液3；所述贮存液3的组成成分为：磷酸盐缓冲溶液：每升含量包括磷酸二氢钾10g、磷酸氢二钾25g、磷酸氢二钠20g和氯化铵2g；硫酸镁溶液11g/L；氯化钙溶液28g/L；氯化铁溶液0.15g/L。本发明的培养基的贮存液3有利于加速大肠杆菌产生酶分解底物,突出的是针对低浓度大肠杆菌检测能缩短检测时间。贮存液3为独立的盐溶液,在0-4℃稳定保存6个月以上,方便实现在线监测。本发明培养基用于检测大肠杆菌的方法,检测结果为大肠杆菌的具体浓度值。(The invention relates to a culture medium and application thereof for detecting escherichia coli, belonging to the technical field of escherichia coli detection, wherein the culture medium comprises an enzyme substrate 1 of 4-methylumbelliferone- β -D-glucuronide, a nutrient 2 of tryptone or peptone and a stock solution 3, wherein the stock solution 3 comprises phosphate buffer solution with the content of 10g of potassium dihydrogen phosphate, 25g of dipotassium hydrogen phosphate, 20g of disodium hydrogen phosphate and 2g of ammonium chloride per liter, 11g/L of magnesium sulfate solution, 28g/L of calcium chloride solution and 0.15g/L of ferric chloride solution, the stock solution 3 of the culture medium is favorable for accelerating the escherichia coli to generate enzyme decomposition substrates, and the detection time can be shortened aiming at low-concentration escherichia coli, the stock solution 3 is an independent salt solution and can be stably stored at 0-4 ℃ for more than 6 months, so that online monitoring can be conveniently realized.)

1. A culture medium comprising: enzyme substrate 1, nutrient 2, and stock solution 3;

the enzyme substrate 1 is 4-methylumbelliferone- β -D-glucuronide (MUG);

the nutrient 2 is tryptone or peptone;

it is characterized in that the preparation method is characterized in that,

the storage solution 3 is a salt solution and comprises the following components:

phosphate buffer solution: the content of each liter of the compound comprises 10g of monopotassium phosphate, 25g of dipotassium phosphate, 20g of disodium phosphate and 2g of ammonium chloride;

magnesium sulfate solution with the content of 11g per liter;

calcium chloride solution with a content of 28g per liter;

ferric chloride solution, 0.15g per liter.

2. The culture medium according to claim 1, comprising:

the enzyme substrate 1 is 4-methylumbelliferone- β -D-glucuronide, and the mass concentration of the enzyme substrate is 0.25-150 mg/L;

the nutrient 2 is tryptone, and the mass concentration of the tryptone is 0-10 g/L;

the dosage of each component of the stock solution 3 is 0.2-5 mL/L.

3. The culture medium according to claim 1, comprising:

the enzyme substrate 1 is 4-methylumbelliferone- β -D-glucuronide (MUG), and the mass concentration of the enzyme substrate is 75 mg/L;

the nutrient 2 is tryptone, and the mass concentration of the tryptone is 10 g/L;

the stock solutions 3 each contained 1 mL/L.

4. The culture medium according to any one of claims 1 to 3, wherein amphotericin B or an extract of Solanum or bile salts is added to the culture medium when fungi or gram-positive bacteria are detected in the actual sample.

5. The medium according to claim 4, wherein the amphotericin B is added to 1L of the medium at 1mg, or the extract of the plant belonging to genus Solanum at 500mg, or the bile salt at 1.5 g.

6. Use of a culture medium according to any one of claims 1 to 3 for the detection of E.

7. Use of the culture medium according to claim 6 for the detection of E.coli, comprising the following steps:

step 1, preparing a culture medium according to a formula;

inoculating escherichia coli into a pure culture medium by aseptic operation, culturing to a stationary phase, diluting to obtain a series of concentration gradients, and preparing a standard curve;

step 3, quantifying the bacterial liquid obtained in the step 2 by using a standard plate counting method (CFU) to obtain the true concentration of the pure cultured escherichia coli;

step 4, inoculating the bacterial liquid obtained in the step 2 into the culture medium prepared in the step 1, and calculating the number of escherichia coli contained in each bacterial liquid according to the CFU quantitative result obtained in the step 3;

step 5, inoculating an actual sample to be tested into the culture medium prepared in the step 1, and adding amphotericin B or solanum plant extracts or bile salts; 6, incubating the culture medium obtained in the step 4 and the step 5 at 35-44 ℃ for 6-19h, measuring fluorescence intensity, exciting 366nm, and reading the peak value of an emission peak at 450 nm;

step 7, drawing a standard curve according to the number of the escherichia coli measured in the step 4 and the emission peak value measured in the step 6; and (5) substituting the peak values measured in the step (5) and the step (6) into a standard curve, and calculating to obtain the concentration of the escherichia coli in the sample to be measured.

8. The use of the culture medium according to claim 7 for detecting Escherichia coli, wherein amphotericin B1mg, or Solanum plant extract 500mg, or bile salt 1.5g is added to 1L of the culture medium in step 5.