plasmid cloning

[ntdna]

i was wondering about plasmid cloning and why some plasmids enter to the cell (EcoRI for exemple) and others don't !!

Edited December 5, 2010 by ntdna

[rasing02]

Do you mean Ecoli Plasmid? Various kinds of plasmid can be transformed into cell via chemical transformation or electroporation and similar process, I hope I am answering the question you addressed? Is this what you asked? if not then sorry..i tired

[ntdna]

Why are some plasmids able to enter cells (ex: EcoRI)? What unique properties do they possess?

 

thank you

Edited December 6, 2010 by ntdna

[CharonY]

Using aggressive methods essentially all plasmids can be transferred into the cell. They just may not propagate (lacking e.g. the proper ori, being lethal to the cell, etc.)

[DeltaScience]

Basically you need the plasmid to encode an essential gene for the microorganism, lets say for instance a plasmid encoding a gene to survive in a medium with an antibiotic, or a gene enconding a protein to degrade an essential substrate for the cell to grow, like lactose, and then keep the transformed microorganism under those conditions, this is like mandatory selection, you're forcing the cells to be under an specific condition in wich they need to use the genes that are present in the plasmid, this way you can guarantee that your cell wont loose the plasmid in the future, this is because it requires more energy for the cell to replicate this DNA.

 

Also you need to take into account different parameters in order to get a succesfull transformed cell, like the size of your plasmid, if its too big it will be harder to get into the cells, and its also easier for the cell to loose it; what kind of genes it poses, if its circular or lineal, etc.

 

I hope this helps to answer your question.

 

greets

Edited January 25, 2011 by DeltaScience