λ Phage phage spot picking experiments
Genetically homologous λ phage protospecies are obtained by "picking" a completely separate phage spot and preserving agar/agarose containing the lysate in a storage solution. The obtained protospecies can be used to prepare plate lysates or liquid cultures of phage, and can also be used for later analysis. The source of this experiment is "Guide to Molecular Cloning Experiments, Third Edition" Translated by Huang Peitang et al.
Operation method
Picking experiments of λ phage phage spots
Principle
Genetically homologous λ phage protospecies are obtained by "picking" a completely separate phage spot and preserving agar/agarose containing the lysate in a storage solution. The obtained protospecies can be used to prepare plate lysates or liquid cultures of the phage, and can also be used for later analysis.
Materials and Instruments
λ Phage Move I. Materials For more product details, please visit Aladdin Scientific website.
Chloroform SM
Borosilicate Pasteur tubes (with rubber bulb) or micropipettes Microcentrifuge tubes or polypropylene tubes
1. Buffers and solutions
Chloroform
SM
2. Specialized equipment
Borosilicate Pasteur tubes (with rubber bulb) or micropipettes
Microcentrifuge tubes or polypropylene tubes (13 mm X 100 mm)
3. Carriers and strains
λ phage, which forms completely separate phage spots on the bacterial moss.
II. Methods
1. Add 1 ml of SM to a microcentrifuge tube or polypropylene centrifuge tube and add a drop (about 50 μl) of chloroform.
Polypropylene centrifuge tubes are used because they are resistant to chloroform.
2. Using a borosilicate Pasteur pipette with a rubber bulb or a micropipette, puncture the selected λ phage phage spot up to the underlying hard agar, and gently aspirate the phage spot into the pipette along with the underlying agar.
Alternatively, phage spots can be picked by gently touching the surface of the top layer of agar/agarose in the center of the selected phage spot with an 8-cm-long wooden swab (also known as a picker swab or cotton swab) or toothpick used for seedling eradication, and then immediately placing the swab into SM/chloroform and vortexing it to dislodge the phage-containing agar/agarose.
3. Wash the agar block from the borosilicate Pasteur pipette, place it in a test tube containing SM/chloroform (prepared in step 1), and leave the test tube capped at room temperature for 1~2 h to dislodge phage particles from the agar block. To aid elution, the test tube can be placed on a shaker and gently rotated. Finally, store the phage suspension at 4℃. 