Are they the same bacteria? Or are you trying to transfer a plasmid between two different types of bacteria?

If they're different types of bacteria, you'd need to make sure that whatever plasmid your resistance cassette is on actually works in that bacteria.

I might be misunderstanding what you want to do/how you want to do it, but if theyre the same bacteria eg E. coli, why can't you just purify the plasmid DNA with midi prep and transform that into your strain of E. coli and then plate on the appropriate selective media? Pretty sure most transformations involve heat shock of competent cells.

I guess what you want to do is kinda unclear. Are you trying to show that bacteria can transfer plasmids between each other and gain resistance, or do you just want to get a specific plasmid into your bacterial strain?

Heat shock increases the efficiency of plasmid DNA diffusion across cell membranes.

For example from bacterium A to bacterium B:

Then, (1) perform miniprep from bacterium A and (2) make competent cells from bacterium B. Then, heat shock the plamsid from (1) into bacterium B competent cells.

Purify the DNA from Bacteria (donar) and be ready with competent cell of target and do transformation using heat shock method. This method is actually more efficient. The transformation efficiency is more when compared to other methods.

Artificial competence can be induced in laboratory procedures that involve making the cell passively permeable to DNA by exposing it to conditions that do not normally occur in nature.[41] Typically the cells are incubated in a solution containing divalent cations (often calcium chloride) under cold conditions, before being exposed to a heat pulse (heat shock). Calcium chloride partially disrupts the cell membrane, which allows the recombinant DNA enter the host cell. Cells that are able to take up the DNA are called competent cells.

It has been found that growth of Gram-negative bacteria in 20 mM Mg reduces the number of protein-to-lipopolysaccharide bonds by increasing the ratio of ionic to covalent bonds, which increases membrane fluidity, facilitating transformation.[42] The role of lipopolysaccharides here are verified from the observation that shorter O-side chains are more effectively transformed – perhaps because of improved DNA accessibility.

The surface of bacteria such as E. coli is negatively charged due to phospholipids and lipopolysaccharides on its cell surface, and the DNA is also negatively charged. One function of the divalent cation therefore would be to shield the charges by coordinating the phosphate groups and other negative charges, thereby allowing a DNA molecule to adhere to the cell surface.

DNA entry into E. coli cells is through channels known as zones of adhesion or Bayer's junction, with a typical cell carrying as many as 400 such zones. Their role was established when cobalamine (which also uses these channels) was found to competitively inhibit DNA uptake. Another type of channel implicated in DNA uptake consists of poly (HB):poly P:Ca. In this poly (HB) is envisioned to wrap around DNA (itself a polyphosphate), and is carried in a shield formed by Ca ions.[42]

It is suggested that exposing the cells to divalent cations in cold condition may also change or weaken the cell surface structure, making it more permeable to DNA. The heat-pulse is thought to create a thermal imbalance across the cell membrane, which forces the DNA to enter the cells through either cell pores or the damaged cell wall.

Electroporation is another method of promoting competence. In this method the cells are briefly shocked with an electric field of 10-20 kV/cm, which is thought to create holes in the cell membrane through which the plasmid DNA may enter. After the electric shock, the holes are rapidly closed by the cell's membrane-repair mechanisms.

@ Michael Daddario

Yes, I am trying to see whether or not the bacteria can transfer plasmids between each other and gain resistance.

I've never heard of enhancement of strain-to-strain transfer by heat-shock, though generally speaking, stress tends to induced genetic transfer mechanisms.

May I ask where you got the idea from?

Andrew Jenkins

@ Andrew Jenkins

I didn't get this idea from any establish work!It's nothing but my own thoughts!

Thank you for replying!

Give it a try and publish it if it works!

Alternatively, try subinhibitory concentrations of nalidixic acid. This will induce the SOS response, which often stimulates plasmid transfer.

Andrew

Wow!

Thanks for the idea @Andrew Jenkins