Reprogramming Life

President Obama is expected to lift the ban on federal fund for embryonic stem cell research soon. However, that does not seem to be the hottest topic these days concerning stem cell research. In 2006, Shinya Yamanaka showed that mouse skin cells could be reprogrammed back into something called induced pluripotent stem (iPS) cells by introducing a handful of cDNAs using retroviral vectors. The process was later repeated in human cells and by other groups including those of Thomson and Melton, sometimes with a slightly different set of inducing cDNAs, or with chemicals or shRNA repressing the repressors of the inducer genes.

The iPS cells are not exactly the same as ES cells, and no animals have been created using iPS cells, but they are close enough to be of great interest to lots of people, particularly for basic research purposes. The method to create iPS by reversing chromosomal changes along differentiation pathways appears to be surprisingly simple, like erasing an old audio tape, there may still be acoustic information left if analyzed by the right equipment, but to most people it is as clean as new. You’d wish a few things in life could be reversed that easily!

For labs that are not already in the stem cell field but feel a need to get their feet wet, then they want reagents that are pre-assembled and pre-tested. Such reagents may include: iPS cultures, iPS inducing viral particles, antibodies to stem cell specific markers, cell assays, and even PCR primer sets (synthesizing hundreds of oligos used in the Yamanaka papers alone will take a lot time and unnecessary costs). That’s where a fast-moving, research-oriented company like Allele comes in. We will bring what we think as starter sets for you, and listen to what you think as needed as we along. The new iPS product line will be launched within weeks, hopefully coinciding with our brand new webpages for all our current product lines!

Tags: c-myc, desease tissue regeneration, differetiation, iPS, Klf4, Lin28, Nanog, Oct3/4, pathways, pluripotent, re-programming, Sox2, stem cells, therapy

What’s In It for Us?

$10 billion will be used to support research projects under the National Institutes of Health over two years, with approximately $1 billion earmarked for cancer research. $7.4 billion will be spent on R01 by the NIH before September 30th, 2010. In the mean time, NSF gets $3 billion for the same period of time.

What do these numbers mean? They would translate into approximately 14,000 RO1 grants at the NIH and nearly double the funding percentage at the NSF. If your lab has grant application(s) that is being hung up on uncertainty, now you have some certainty that they should be funded. If you have been having trouble gathering inner strength to go through the grant writing process, maybe it is time to feel good and potentially rewarding about coming back to the writing table.

Good luck!

P.S. Come back to check Allele Biotech’s new iPS product line, and blogs on induced stem cell research.

Tags: biology department jobs, grants, NIH NSF budget, research dollars, research market

Effective Use of Resources in Difficult Times

In scientific research, there is a tendency to have everything done in our own lab just so that you can say so, after all, scientific credit is the core criterion researchers are evaluated on. You say wait a minute, don’t we always encourage exchange of materials and COLLABORATION on projects? Sure, but not often enough to make “encouragement” unnecessary. Many “collaborations” are more like sharing of materials with conditions.

In business, collaboration is more in the form of OUTSOURCING or CO-DEVELOPMENT (sometimes through licensing), because doing everything by one’s own employees just doesn’t make much financial sense even for the mega-sized, we-have-everything type of companies. One friend of ours working at a Johnson & Johnson site once told us that a line of research using gene silencing technologies was debated but never moved forward because the lack of confidence in expertise: we are not expert on RNAi, how do we trust our own data? For most biotech and early-stage pharma companies, hiring an expert to do a task brings about too much uncertainty, not to mention cost efficiency.

“Having the expert do it” by outsourcing is somewhat more acceptable to the industry than the academia because the “We are the experts” mentality is more dominant in the latter. Heck, if we don’t believe “We are the experts” in our own field of research, then why do we even do it in the first place? In business though, who is the expert is not something one fights for if the end product or contribution to profit is not made.

The current economic conditions caused many large biotech and pharma companies to lay off thousands upon thousands of employees, in one case of Pfizer layoff, scientist positions were particularly targeted for elimination. Life goes on. Economic downturns are also opportunities for becoming lean and mean, using ways of doing things with much improved efficiency and productivity such as outsourcing, and finding new areas for long term growth.

Tags: assay development, cell culture, co-development, collaboration, molecular biology services, outsourcing, project, protein expression, stem cell development, stem cell reagents, stem cell services, tasks

$6.5 billion more to the NIH

On Feb 3^rd the U.S. Senate agreed to an amendment that would provide an additional $6.5 billion to the National Institutes of Health (NIH) for biomedical research as part of the American Recovery and Reinvestment Act of 2009.

Last week, the House of Representatives and the Senate Appropriations Committee voted for the American Recovery and Reinvestment Act, which included $3.5 billion in supplemental funding for the NIH. The total stimulus package, still being amended in the Senate, is now over $900 billion.

The Senate is still debating the bill and being encouraged to finish it as early as this Friday.

Tags: $6.5 billion, Allele Biotech, billion, biomedical research, budget, funding, grant, NIH, R&D, science budget

Getting the most from fluorescent proteins, Part 2

On Feb 3^rd in our previous blog entry on fluorescent proteins, we discussed some basic tips on setting yourself up for success with fluorescent protein based experiments. Here are some more ideas to help boost your imaging success:

1.    Know your background.

All cells contain endogenous fluorescent materials which can confound image interpretation, especially when your fluorescent protein signal is weak.  Make sure you’re familiar with the autofluorescence of your cell type before starting your FP experiments: take some images of non- expressing control cells using the same filters and excitation wavelength as you plan to use for your FP imaging.

Keep in mind that for mammalian cells, autofluorescence is confined mainly to the blue and green regions of the visual spectrum, while in other organisms (e.g. plants, yeast, and bacteria), some cell types may contain fluorescent compounds in other regions of the spectrum. For any given species and cell type, there is likely to be a wavelength “window” with the least autofluorescence; try to choose a fluorescent protein in this wavelength range for maximum signal above background.

2.    Sometimes two (or more) FPs are better than one.

If you are having trouble obtaining sufficient fluorescent signal from a fluorescent protein fusion construct, consider adding an extra copy of the fluorescent protein to boost your brightness.  While this is not recommended unless all else has failed, for low-abundance proteins it can substantially increase the likelihood of detection.  It is possible to create a functional fusion of two or more copies of fluorescent protein in many cases, although the larger size of such a tag increases the chances of mislocalization, so proper controls and validation are essential if you use this technique.  Also, remember that it is generally difficult to use PCR to amplify tandem copies of any gene, including FPs, so restriction-based subcloning is the most reliable way to create multi-copy FP tags.

3.    The best fluorescent proteins don’t stick together!

Truly monomeric fluorescent proteins make the best fusion tags, since they don’t produce localization artifacts due to multimerization. Even weak dimers, such as EGFP and its derivatives, can cause trouble if your fusion protein is at high concentration or in a confined space like a membrane or vesicle.

Are you still using your old EGFP fusion constructs?  If so, make sure to validate your localization results by other methods, or switch to a truly monomeric FP such as mTFP1 or mWasabi.  If you prefer to keep your original constructs, note that any Aequorea GFP-derived FP can be made completely monomeric by adding the A206K mutation.

One final warning — many commercially available FPs that were initially advertised as being monomeric later turned out to be dimers!  With any new FP you try, validate your results before making your conclusions.

Tags: cell marker, cellular, cellular localization, fluorescence, fluorescent protein, FRET, GFP, imaging, mTFP1, mWasabi