Hi Nauman,

Typically you would only want to insert a single gene into any given expression vector.  There are vectors that would allow for 2 genes, provided you are looking to make a fusion protein using an IRES (internal ribosome entry site).  IF you are just looking to catalog and store your genes of interest for future work, you could potentially insert 2 or 3 into a vector provided the restriction sites work for you.  

There is typically an "upper limit" to the size that you can transform into bacteria.  There are certain strains of bacteria that allow for larger constructs 10KB+ (XL10Gold and others).

Hi Nauman,

Typically you would only want to insert a single gene into any given expression vector.  There are vectors that would allow for 2 genes, provided you are looking to make a fusion protein using an IRES (internal ribosome entry site).  IF you are just looking to catalog and store your genes of interest for future work, you could potentially insert 2 or 3 into a vector provided the restriction sites work for you.  

There is typically an "upper limit" to the size that you can transform into bacteria.  There are certain strains of bacteria that allow for larger constructs 10KB+ (XL10Gold and others).

Hi,

Agree with above answer...

If you want to express your gene then you can only insert single gene in MCS (multiple cloning site) of expression vector.

I have fused two and three genes in single expression vector to express them into single chain in form of fusion proteins. But if you want to express differently like your gene of interest and GFP like protein then you should search for dual or multi promoter vectors with different position and MCS but it is very tricky.

I have successfully transfected 8-9kb plasmid into routine DH5alpha, but by using commercially available complement cells you can achieve 10KB+.

Hope this will help.

Thankx

dear Nauman

maybe you can find interesting the following article:

Scheich C, Kümmel D, Soumailakakis D, Heinemann U, Büssow K. Vectors for

co-expression of an unrestricted number of proteins. Nucleic Acids Res.

2007;35(6):e43. Epub 2007 Feb 20.

of course the lenght of the genes isi mportant because as Barry suggest there is a limitation size in the plasmid that you can trasfrom in the E.coli, so if you have small genes the number is higher respect than big genes.

good luck