PCR validation of basic biological experiments in [life sciences]

Basic biological experiments PCR verification

Experiment step one

Add... Into the centrifuge tube 7.5μL Master Mix solution ,5.5μL distilled water , Each primer 0.5μL μL as well as 1μL Templates （ Of various plants DNA）.

Experiment step two

1 95℃ 3min Heating up
2 95℃ denaturation 30 s
3 58℃ perhaps 60 ℃ Anneal 30s
4 72℃ extend 45s
5 72℃ extend 45s end
choice Files, Click on program, Click New . And then click Header, If it is done PCR Words ,Heaterd Lid Temperature selection 105℃. Don't worry about the rest . The actual translation time setting of the picture may be inconsistent with the text steps , The text steps shall prevail .

Experiment step three Dispensing

Waiting for the PCR The result of the process , Prepare the glue first . weighing 0.3g agarose , Join in 20ml TAE solution , Mix and place in the microwave 2min after When the solution is transparent Join in 1μmol/L Nucleic acid dye .（ Gloves must be worn during the experiment ）. The agar concentration depends on DNA Segment size , The clip is big , Low concentration ,1000bp The concentration is about 1.5%. Fragment small , High concentration ,200bp The concentration is about 2-3%. Put it into the gel bath . According to need , Select plug-in boards of different sizes as shown in the following figure , The plug-in board has different sizes . Large plug-in boards can be used for large-scale experiments . wait for 15 Minutes after solidification, transfer to electrophoresis instrument .

Experiment step four electrophoresis

take PCR Good response DNA 7.0μL Insert into the sample hole , Put... On the far right Marker5000（ Depending on the DNA Choose different lengths Marker）. voltage U=140V, reaction time =15 minute . Then start electrophoresis ！

Experimental step 5 Glue running

electrophoresis 15 Minutes later , After taking out the gel, carefully move it into the electrophoresis instrument for reading .

Click on the top left corner to take photos , Then click the camera icon in the lower left foot , The analysis begins , Get electrophoresis results . according to DNA MAKER contrast DNA Fragment size . This is the end of the experiment ！