We've posted the manuscript on the public arXiv.org server. You can download the full pdf, including all the supplementary data, at http://arxiv.org/abs/1201.6643.
7 hours ago in The Phytophactor
Distribution on the Internet may be considered prior publication and may compromise the originality of the paper as a submission to Science. Please contact the editors with questions regarding allowable postings.Has anyone had direct experience with this? I think I'd better send out a tweet...
The amount of phosphate in the medium used by Wolfe-Simon et al for their published growth analysis is indeed uncertain. Their ICP-MS analysis found that most of their media preparations contained 3-4 µM phosphorus, but one batch contained <0.3 µM and a solution containing only the AML60-medium salts had 7.8 µM. Because we don't know which batch was used for the results in their Figure 1, 3-4 µM is a good estimate of the phosphorus contamination, but the actual amount could have been substantially lower or higher.
My cells did grow in medium with no added phosphorus*, to about 5 x 10^6 cells/ml. This is about 1/4 of the density reached by GFAJ-1 in Wolfe-Simon et al's '-P/+As' medium. Adding 3 µM phosphorus to my medium increased GFAJ-1 growth fourfold, to the same density as reported in Wolfe-Simon et al's experiments. Simple algebra thus suggests that my unsupplemented medium contained about 1 µM phosphorus. The correspondence of the cell densities reached in my supplemented (3 µM) and their unsupplemented medium supports the estimate of 3-4 µM contaminating phosphorus in their medium.
My cells, like theirs, were clearly phosphorus-limited, because they grew to much higher densities when additional phosphorus was provided (see my recent RRResearch post and their Fig. 1).
I think this is the best that can be done, since Wolfe-Simon et al. apparently did not keep good enough records to determine the actual phosphorus concentration of the medium they used for their reported experiments.
Hope this helps,
*The initial growth problem was not due to a lack of phosphorus but to the need for an amino acid, which I solved by supplementing the medium with a small amount of glutamate.
In the second paragraph they seem to be first saying they'll recommend my already-published paper to their editors (for the editors to do what, read it with admiration?), and then asking me to add a bit of new material to it and submit it to them for publication. This reeks of self-plagiarization. But in the next sentence they ask for other papers instead.Dear Rosemary J. Redfield,
This is Scientific & Academic Publishing, USA. Nice to get your information from the journal PLOS Pathogens and also happy to pass on our regards to you from the editorial department of SAP.
We've finished reading the abstract of your paper Transformation of Natural Genetic Variation into Haemophilus Influenzae Genomes and will recommend it to our editors. If you are interested in our journals and want to publish it on our journals, please extend this paper and describe your latest research achievements and send it to us by our online submission system (http://www.manuscriptsystem.com). All manuscripts submitted will be considered for publication.
If this paper has been published, we also welcome you to submit other papers to us.
Welcome to visit our website at http://www.sapub.org.
im curious about how much time the DNA you prepared has spent in solution since you prepped it — supposing there were C-Ar bonds in the DNA at the time of lysis, has so much time passed now that, given the hydrolysis rates that were experimentally determined and recently reported in this JACS communication (offensive link deleted), I'm worried that theres no hope of seeing positive signal even if there was one to begin with. Any thoughts about this issue? Perhaps keeping the DNA stored as a dry pellet and avoiding any encounters with water until the last minute would be the way to go.First, a few corrections: Arsenic is As, not Ar. The DNA backbone bonds are diester bonds, so the As is bound to oxygen (O), not carbon (C). It's of course not possible to purify DNA while avoiding any encounters with water.
Since the cell’s interior is aqueous, then it would seem reasonable that there has to be something preventing the spontaneous hydrolysis in vivo. As you point out, some form of stabilizing protein would seem the most likely candidate. If this protein is going to work, then it could (for the purposes of this argument) remain attached throughout the purification process. If so, then you might find As in the purified DNA, and should also find traces of protein. I don’t know if this would be enough to alter the 260:280 ratio, but DNAse digestion followed by SDS-PAGE might detect something. Definitely a hotdog experiment, but once the DNA is purified, not that difficult.SDS-phenol extractions are pretty harsh; for a protein to remain with the DNA it would need to have been covalently bound, which would certainly have complicated such cellular processes as DNA replication and transcription. And if the DNA contained a significant amount of As, the DNA would then contain enough protein that it wouldn't migrate properly in the gel but would stick as a blob of gunk in the well at the top of the gel. It also would band differently in a CsCl gradient.
HI Dr.Rosie Redfield,
Here's a chance to represent UBC's Zoology Science Department.
The annual Science Week Events Committee , organized by the Science Undergraduate Society, would like to invite you to join our event on Thursday, January 26th, 2012 at 12:30-1:45.
As you may know, Science Week, is a week-long (form January 23rd - January 27th), multi-events celebration which allows students to show off their UBC pride while rewarding them for their first term achievements. So far we have jell-O wrestling, jeopardy, and a scavenger hunt. Although, we realized something was missing...(*1)
This year, we have added a new event to our venue, called the "Professor Pageant". UBC Professors, like yourself, will be participating in the pageant and showcasing their popularity, attitude and talents (*2). Only one competitor will come out on top, however, each participant will go home with a special customized award.
We would like to invite you to join us because you are deemed awesome (yes, awesome) by an overwhelming consensus among students (*3). This is an opportunity that you do not want to miss! (*4)
We would love to hear from you and provide more details. Also, if you would like to suggest any of your colleagues (open to all faculties), please provide their name so we could can invite them accordingly!
(Names withheld to protect the naive.)
The green data: 6778 ng of DNA (89% of the total DNA recovered) is in four fractions with a total volume of 300 µl (37% of the volume recovered). This means that the concentrations of soluble contaminants not bound to the DNA will have been reduced to about 40% of what they were.
The red data: 5135.3 ng of DNA (68% of the total DNA recovered) is in two fractions with a total volume of 310 µl (17% of the total volume recovered). This means that the concentration of soluble contaminants will have been reduced to about 25%.