Plasmid extraction kit and plasmid extraction method
A kit and plasmid technology, used in biochemical equipment and methods, microbial determination/inspection, DNA preparation, etc., can solve problems such as inability to recover plasmids for proteins and plasmids, residual protein cleavage, water pollution, etc. The effect of pressure, less environmental pollution, and reduced consumption
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
Embodiment 1
[0036] Example 1 Construction of Plasmid Extraction Kit
[0037] The plasmid extraction kit of this embodiment includes solution 1, solution 2, solution 3, solution 4, solution 5 and flocculation powder. The solution 1 is double distilled water, the solution 2 is 0.21M NaOH solution, the solution 3 is a concentrated salt solution containing 1.6M potassium acetate, 1M acetic acid and 0.11M calcium chloride, and the solution 4 is 70 % ethanol solution, the solution 5 is an RNA clearing solution containing 10mM Tris-HCl, 1mM EDTA and 10μg / ml RNase, and the flocculation powder is a mixture of NaOH and SDS in a mass ratio of 1:6.
[0038] Wherein, solution 2, solution 3, solution 4 and solution 5 were all prepared with double distilled water. The reagents used are above the analytical grade. Both solution 3 and solution 5 were stored at 4°C, and other solutions were stored at room temperature.
[0039] Solution 5 was used as the RNA clearing solution. First, a mixture of Tris-HC...
Embodiment 2
[0040] Example 2 Plasmid mini-extraction method
[0041] 1. Cultivate Escherichia coli with 1ml~25ml LB liquid medium containing 1μl / ml ampicillin for 16~24 hours, centrifuge at 2100g for 10min, discard the supernatant, resuspend with 1ml deionized water and transfer to a 2ml centrifuge tube , centrifuge at 16000g for 1min and discard the supernatant;
[0042] 2. Add the reagents according to Table 1:
[0043] Table 1 The amount of small extraction reagents
[0044] The volume of the culture solution Solution 1 Solution 2 Solution 3 Flocculating powder Solution 5 Solution 4 Centrifuge tube 11ml~25ml 250μl 500μl 750μl 0.5g 250μl 500μl 2ml centrifuge tube 1ml~10ml 100μl 200μl 300μl 0.2g 100μl 500μl 1.5ml centrifuge tube
[0045] 3. Add solution 1 to the cell pellet according to Table 1, and vortex to resuspend the cell evenly;
[0046] 4. Add solution 2, invert 6 to 10 times to mix well, and let stand at room tempe...
Embodiment 3
[0059] Example 3 Plasmid extraction method
[0060] 1. Centrifuge the bacterial solution cultured at 37°C into a 50ml centrifuge tube (less than 500ml bacterial solution), discard the supernatant;
[0061] 2. Add the reagents according to Table 2:
[0062] Table 2 The dosage of large extraction reagents
[0063] The volume of the culture solution liquid 1 Solution 2 Solution 3 Solution 5 Solution 4 Flocculating powder 26～100ml 1ml 2ml 3ml 800μl 1ml 2g 101～300ml 3ml 6ml 9ml 800μl 1ml 6g 301～500ml 5ml 10ml 15ml 800μl 1ml 10g
[0064] 3. Add solution 1 to resuspend bacteria according to Table 2, add solution 2 and invert 10 times to mix well, and let stand at room temperature for 5 minutes;
[0065] 4. Add solution 3 according to Table 2, shake vigorously up and down 10 times, centrifuge at 2100g at 4°C for 5min, slowly pour the supernatant into a new 50ml centrifuge tube, avoid pouring large precipitates;
[0066] 5. ...
PUM
Login to View More Abstract
Description
Claims
Application Information
Login to View More