News

The BGSC is pleased to announce the availability of four new integration vectors, pXFP_bglStar, to generate low noise fluorescent promoter fusions to insert into the bglS locus in Bacillus subtilis. They are available in the catalog under the BGSC IDs ECE793-ECE796.

These will add on to the pXFP_Star vector suite (ECE295-ECE297) which buffer against spurious transcription by having a terminator.

Thank you to Dr. Ilka Bischofs-Pfeifer of the Max Planck Institute for Terrestrial Biology for donating these vectors.

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Thanks to the generosity of many researchers in the field of Bacillus research, we have accepted donations for several new strains over the past year. We are happy to announce that those strains have now be added to our online catalog and are available for order. Please be on the lookout for more announcements as we introduce these new strains.

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Thank you all for your patience during our transition! We're scaling down our COVID-19 testing operation and that has allowed us much needed time to catch-up. We recently completed a transition from a manual credit card console to an online credit card payment processing system. This allows us to send you a direct payment link through email, providing you the flexibility to complete payment at your convenience. We're also working hard on a new website with integrated payment processing which we hope to launch this fall.

Please direct all inquiries to our new email address - asc-microbioservice@osu.edu. Our team will follow-up with you by email and can schedule calls as needed. Thank you!

Jeff

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As many of you have heard, I am retiring from Ohio State on 28 February and will be working as an independent consultant. Beginning 1 March, the BGSC will have a new collection manager, Jeff Jahnes. Jeff came to Ohio State in 2013, first serving as Laboratory Supervisor for the Department of Microbiology, then in 2019 assuming the directorship for the Applied Microbiology Services Lab (AMSL) at Ohio State's Infectious Diseases Institute. As the pandemic unfolded, Jeff was charged with developing and managing the COVID-19 testing facility that serves Ohio State's 50,000+ students. He's done a fantastic job in these positions, so I feel fortunate that he has agreed to direct the BGSC. Strain maintenance and distribution will continue uninterrupted during the transition, but please be patient: Jeff and his team do face a bit of a learning curve as they implement and eventually improve upon our systems. Jeff's contact information is below:

email: ASC-microbioservice@osu.edu

office: +1 614-292-3490

I want to thank the Bacillus research community for all the positive interactions and teamwork over the last 35 years. Our field seems to attract talented, interesting, creative people who value collaboration. I hope to stay active in science through consulting, editing, and writing. If you want to get in touch with me, my OSU email address should remain active (zeigler.1@osu.edu). Best wishes for a safe and productive 2021--and beyond!

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Five copies of the FreeGenes "Bacillus subtilis genes (E. coli codon optimized) expression library" are available. As described on the FreeGenes website, "261 essential genes from Bacillus subtilis, codon optimized for Escherichia coli in MoClo-compatible high-copy ampicillin resistance backbones. Shipped in Escherichia coli Top10 in three 96 well plates." I will distribute them on a first come, first served basis this week only. (Of course, you can also obtain them from FreeGenes after the week is up.) Please contact me directly for details at zeigler.1@osu.edu.

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Forwarded message from Emma Denham and Matt Cabeen:

Dear Bacillus Global Community,

We hope that you and your family are well during this time.

As we continue through these unprecedented times and to fill the gap until we can be together at in person conferences. We are pleased to announce Subtillery 2021 - a virtual meeting for all things Bacillus. This will take place June 14th-18th on Zoom.

Sessions will run for approximately 4 hours, as with Subtillery 2020 with the following start timings.

6:45am Los Angeles, USA

8:45am Chicago, USA

9:45am Washington DC, USA

10:45am Brasilia, Brazil

2:45pm London, UK

3:45pm Paris, France; Berlin, Germany

7:15pm New Delhi, India

9:45pm Beijing, China

10:45pm Tokyo, Japan

11:45pm Canberra, Australia

The conference website can be found here: https://cabeenlab.okstate.edu/subtillery-2021

We are very grateful to BACIP for sponsoring the speaker awards for this meeting.

We will be accepting abstract submissions for talks from March 1st. Abstract submission will close May 7th. Decisions will be sent out on the 21st May. Registration will also open on the 1st March.

Best wishes,

Emma and Matt

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From Michel Gohar:

"The next BACT meeting, initially planned on September 2020, had been rescheduled to take place in Paris in April this year.

"Unfortunately, since the Covid19 pandemic is not expected to be fixed before summer in Europe, the physical meeting will be replaced by an online meeting, from April 26 to April 28 2021.

"Presentations from PhD students and postdoctoral fellows will be selected in priority for this meeting. Registrations and abstract submissions will open on February 3rd, on the BACT2021 website https://colloque.inrae.fr/bact2021.

"The regular meeting in Paris is not cancelled, but postponed to April 2022."

For those new to the field, the International Conference on Bacillus anthracis, B. cereus and B. thuringiensis (BACT) is a wonderful opportunity to learn from and network with other researchers who focus on species in the Bacillus Cereus Group. This would be a great opportunity for grad students and post docs to gain some visibility in this research community.

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On March 1, 2021, the Bacillus Genetic Stock Center will transfer to a new earnings unit at The Ohio State University. In the next few weeks I will introduce a new collection manager. The website will also change, and there will be an improved ordering process. The stocks will still be available, and prices will remain stable the rest of the fiscal year. We intend the BGSC to maintain and distribute its stocks in perpetuity!

Because we will be unable to transfer legacy commitments to the new earnings unit, one practical consequence is that existing bulk purchase plans should be used up by 28 February. Please plan accordingly, and let me know if this requirement causes you any difficulties.

I will soon be retiring from Ohio State after 35 years of service and will begin a career as an independent consultant. You will still be able to reach me at zeigler.1@osu.edu. Please feel free stay in touch. It has been a pleasure serving all of you!

Watch this spot for more details soon.

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After a nearly 11-week lockdown, the BGSC will be reopening on Wednesday, 3 June 2020. Initally we will be onsite for 20 hours a week. We will begin shipping orders right away. There are a large number of backorders, so it may take us a few weeks to get totally caught up. Please do not hesitate to begin placing orders. We will process them just as soon as we possibly can!

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The BGSC remains under lockdown conditions. We hope to resume operations on or around June 1, 2020. We pass on updates as they become available.

In the meantime, we are happy to announce that Prahathees Eswara of the University of South Florida is organizing a virual conference called Subtillery, with the stated aim of "distilling and understanding the biology of Bacillus subtilis and related organisms." The plan is to have nearly 40 twenty-minute trainee talks presented on Zoom between June 8-12 (10am-1:30pm US Eastern Time). A registration form can be found at Subtillery - Registration

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As most of your probably realize, the Bacillus Genetic Stock Center is hosted on campus at The Ohio State University in Columbus, Ohio. Like most educational institutions around the US and worldwide, Ohio State is grappling with ways to mitigate the severity of the current coronavirus pandemic. The situation is fluid and changing rapidly. Currently, face-to-face classroom instruction has been suspended, and university personnel who are able to work remotely are required to do so. There is a high likelihood that other measures, including a lock down of laboratories, will be mandated in the near future. For this reason, the BGSC will have limited functionality for an extended time. It will be possible to reach us by email (zeigler.1@osu.edu) and by telephone (1-614-292-5550) during regular working hours, 8 am - 5 pm EDT (UTC-04:00). If you are open to receive shipments and are able to make purchases, it may be possible for us to distribute strains. However, I cannot guarantee our continued ability to do so. Please inquire by email before attempting to make a purchase. Thanks for your understanding. Wishing you health and happiness in the days ahead, Daniel

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What makes Bacillus subtilis 168 such a powerful model system? Many factors: a world-wide community of investigators and a 60-year history of focused enquiry; a carefully annotated genome sequence with associated proteomic and transcriptomic data; a suite of useful curated databases making all of this information discoverable and accessible. But what sets B. subtilis apart from most other microbial model systems is a genetic toolbox of great sophistication and variety. One important set of tools are designed to introduce marker-free mutations to the B. subtilis genome. Several technologies are available, including Cre-lox marker loop-out and CRISPR methodologies. But an older technique--allele replacement by temperature-sensitive plasmids--remains highly useful. And the method is potentially extensible to any Gram-positive bacterium with an available plasmid-transformation system, whether by natural competence of by physical methods such as electroporation.

One such vector is pminiMAD2 (also called simply pminiMAD in the research literature). Originally constructed by Patrick and Kearns (2008), this shuttle vector replicates normally in E. coli with selection for ampicillin resistance and in temperature sensitive fashion in Bacillus, where selection is for erythromycin resistance. In Gram-positive hosts, plasmid replication is permissive at normal room temperature but restricted at 37°C. In practice, one simply inserts a fragment from the target chromosome, altered with either a point mutation, deletion, or insertion. The plasmid construct is introduced into the host by transformation, and then selection is maintained as the temperature is raised to 37°C. Single crossovers produce Campbell-type insertion events, where the entire vector is integrated into the chromosome at the target locus, flanked on either side by a normal and mutated copy of the insert. Transformants are next cultured at the permissive temperature in the absence of selection. Several generations of growth, usually at room temperature overnight, allow the plasmid to excise from the genome by hommologous recombination. In many cells, the plasmid will be cured spontaneously, leaving behind either a wild type or mutated form of the target locus in the chromosome. A simple screening step by PCR and sequencing can identify the desired mutant.

The PubMed Central database lists over 40 publications that use pminiMAD2 (or pminiMAD). At least 10 were published during the last two years; they are listed in the article citations below. All describe work in B. subtilis 168 or its wild type ancestor NCIB 3610 with one exception. Spacapan et al. (2018) used pminiMAD2 to introduce a marker-free deletion into an environmental isolate of B. subtilis, PS216 (BGSC accession 3A36). In principle, however, the vector could be used with any mesophilic isolate from Bacillus or related genera.

We thank the Dan Kearns laboratory at Indiana University for donating pminiMAD2 to the BGSC. It is available in an E. coli host as our catalog number ECE765.

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