Co-infection with Baculovirus Vectors for Multiple Protein Expression

A question that comes up from time to time is the use of co-infection with baculovirus vectors for multiple protein expression in insect cells.  The need arises if you have produced a number of baculovirus vectors that each synthesise a component of a multi-subunit complex, such as an enzyme or a virus like particle.  Intuitively, you know that it is probably necessary to add many virus particles per cell to

Baculovirus Transfer Vectors Compatible with flashBAC™

A question we are asked from time to time is to identify baculovirus transfer vectors compatible with our flashBAC™ system. We obviously nominate our own extensive range that is available but frequently someone has an old plasmid from years ago that they want to reuse. Originally they may have paired it with a linear DNA system such as BaculoGOLD or BacPAK6 or even wild type DNA where recombinant virus selection

Variable Expression of Multi-Component Protein Complexes

Variable expression of multi-component protein complexes is increasingly necessary as more sophisticated projects are attempted.  But how can this be achieved?  Most of our expression work is based on the baculovirus system, in particular our flashBAC™ vectors that permit easy recombinant virus generation in a one-step process.  These insect-specific viruses offer an ideal platform for producing a wide range of recombinant proteins including nuclear, cytoplasmic, membrane-bound and secreted examples. But

Intermediate scale recombinant protein production

Between initial tests and larger scale up you may want to conduct intermediate scale recombinant protein production using baculovirus vectors.  If you are working with several constructs then this can begin to get expensive of cell culture vessels such as the ubiquitous 125ml conical flasks, in which you can amplify 25-30mls of virus-infected cells.  Space on shakers can also be at a premium in some labs. To address both the

Expression Systems insect cell culture media

OET is pleased to offer a wide range of Expression Systems Insect cell culture media, from the very popular ESF 921, to specialized insect transfection media. Expression Systems’ media are easy to adapt your cells into and the insect specific media gives great protein yield, when compared to rival media.   ESF921 insect culture media OET use ESF 921 for all in-house work, due to it’s high performance record, whilst

Adapting insect cells to a new growth medium?

Have you ever had the experience of adapting insect cells to a new growth medium?  We recently had to do this for a client project because they had used a particular medium previously and were keen to maintain the same conditions to continue the work with us. This process made us realise that switching to a new serum-free insect cell culture medium can result in short term stress responses in

Baculovirus-expressed secreted recombinant proteins

A question that often comes up from clients is what is the best signal peptide to use for baculovirus-expressed secreted recombinant proteins?  While we have discussed the best flashBAC™ variant to use for the production of secreted proteins in a previous blog, we have never considered the role of the signal peptide. In fact, while writing this blog, we realised that there was much that we didn’t know, so will

baculoQUANT titration of recombinant baculoviruses

There are many methods available for the titration of recombinant baculoviruses.  One of the most convenient is the use of the quantitative polymerase chain reaction (QPCR), which assesses how much virus DNA is present in a freshly-amplified stock.  This is compared with a standard curve derived from a control virus stock to give an infectious titre expressed as Q plaque forming units (pfu) per millilitre.  OET Ltd market the baculoQUANT™

Sequence errors in your gene for expression Part II?

Sequence errors in your gene for expression Part II follows on from an earlier blog where we discussed a problem we were having producing a particular protein after synthesizing a gene based on very old sequence data. The baculovirus we made containing this synthetic gene produced very low levels of recombinant protein that were hardly visible after immunoblot analysis. This wasn’t a good start to a project where a purified

Sequence errors in your gene for expression? (Parts 1 + II)

If you don’t achieve the levels of protein expression expected in your project, have you ever considered that there may be sequence errors in your gene?  This applies to all expression systems, not just those based on baculoviruses.  We should be clear that we are not talking about sequence errors in your gene after it has been synthesized by an out sourcing company.  Quality control by such companies is such

1 2 3 7