Know more

About cookies

What is a "cookie"?

A "cookie" is a piece of information, usually small and identified by a name, which may be sent to your browser by a website you are visiting. Your web browser will store it for a period of time, and send it back to the web server each time you log on again.

Different types of cookies are placed on the sites:

  • Cookies strictly necessary for the proper functioning of the site
  • Cookies deposited by third party sites to improve the interactivity of the site, to collect statistics

Learn more about cookies and how they work

The different types of cookies used on this site

Cookies strictly necessary for the site to function

These cookies allow the main services of the site to function optimally. You can technically block them using your browser settings but your experience on the site may be degraded.

Furthermore, you have the possibility of opposing the use of audience measurement tracers strictly necessary for the functioning and current administration of the website in the cookie management window accessible via the link located in the footer of the site.

Technical cookies

Name of the cookie


Shelf life

CAS and PHP session cookies

Login credentials, session security



Saving your cookie consent choices

12 months

Audience measurement cookies (AT Internet)

Name of the cookie


Shelf life


Trace the visitor's route in order to establish visit statistics.

13 months


Store the anonymous ID of the visitor who starts the first time he visits the site

13 months


Identify the numbers (unique identifiers of a site) seen by the visitor and store the visitor's identifiers.

13 months

About the AT Internet audience measurement tool :

AT Internet's audience measurement tool Analytics is deployed on this site in order to obtain information on visitors' navigation and to improve its use.

The French data protection authority (CNIL) has granted an exemption to AT Internet's Web Analytics cookie. This tool is thus exempt from the collection of the Internet user's consent with regard to the deposit of analytics cookies. However, you can refuse the deposit of these cookies via the cookie management panel.

Good to know:

  • The data collected are not cross-checked with other processing operations
  • The deposited cookie is only used to produce anonymous statistics
  • The cookie does not allow the user's navigation on other sites to be tracked.

Third party cookies to improve the interactivity of the site

This site relies on certain services provided by third parties which allow :

  • to offer interactive content;
  • improve usability and facilitate the sharing of content on social networks;
  • view videos and animated presentations directly on our website;
  • protect form entries from robots;
  • monitor the performance of the site.

These third parties will collect and use your browsing data for their own purposes.

How to accept or reject cookies

When you start browsing an eZpublish site, the appearance of the "cookies" banner allows you to accept or refuse all the cookies we use. This banner will be displayed as long as you have not made a choice, even if you are browsing on another page of the site.

You can change your choices at any time by clicking on the "Cookie Management" link.

You can manage these cookies in your browser. Here are the procedures to follow: Firefox; Chrome; Explorer; Safari; Opera

For more information about the cookies we use, you can contact INRAE's Data Protection Officer by email at or by post at :


24, chemin de Borde Rouge -Auzeville - CS52627 31326 Castanet Tolosan cedex - France

Last update: May 2021

Menu Logo Principal Logo UMR 1332 BFP logo partenaire U-BDX

Home page

UMR 1332 Biologie du Fruit et Pathologie

Wednesday, june 29, 2022 - Thesis defense - Gabrielle GUESDON

Illustration Gabrielle Guesdon
"Development of in-yeast genome cloning methods for the construction of semi-synthetic Bacillus subtilis-derived chassis strains"
Gabrielle GUESDON

Team Mollicutes

Speciality : Microbiology-Immunology

Wednesday, june 29, 2022 - 2:00 p.m. - ISVV Amphitheater - Campus INRAE Villenave d'Ornon

Abstract :

One of the major challenges in the synthetic biology (BS) field, is to provide new solutions to global issues (therapeutic/sanitary or climatic), in particular through the construction of useful, efficient and environmentally friendly production strains.
The well-characterized, non-pathogenic, Gram+ bacterium Bacillus subtilis (Bsu), is widely used in industry as a biotechnological workhorse. Recent studies have established that mutant strains with modified genomes are able to produce larger amounts of recombinant proteins. This suggests that the production of rationally designed Bsu chassis could be an important step in the improvement of valuable strains for industrial purposes.
This work was performed within the Bacillus 2.0's ANR project, which aims at applying SB tools for Bsu, and at developing an effective pipeline for the high-throughput construction of versatile Bsu chassis strains. Selected SB technologies for the pipeline include (i) the synthetic genome design, (ii) the in-yeast DNA assembly methods using Saccharomyces cerevisiae, (iii) the from-yeast whole genome isolation and transplantation (GT) to a recipient bacteria cell and, (iv) the characterization of recombinant strains.

The objectives of this thesis were to ensure the feasibility of these methods using a Gram+ bacterium, by showing, in particular, that it was possible to clone and maintain in S. cerevisiae the genome of a minimal Bsu strain, MPG192 (2.86 Mbp) and to modify it using the large repertoire of yeast genetic tools. Our first attempts to clone the entire Bsu genome into yeast using already described methods failed. Using a TAR-Cloning approach, we then attempted to clone large DNA fragments obtained by restriction of the Bsu genome. In a first experiment, five out of seven fragments were cloned. Difficulties to clone the largest fragment (1.50 Mbp), are presumably related to its size, and/or the lack of ARS elements. Concerning the other fragment, several factors have been proposed to explain the cloning failure: again, an insufficient number of ARS elements, but also, the presence of many repeated sequences (7 ribosomal operons), and/or the deleterious expression of these genes. Finally with other experiments, the whole 2.86 Mb genome was cloned in 21 pieces ranging from 6 kbp to 515 kbp. As TAR-Cloning imposes constraints in the choice of restriction sites, a new cloning method, called CReasPy-Fusion, was developed. This method allows the simultaneous cloning and engineering of mega-sized genome in yeast using the CRISPR-Cas9 system, after direct bacterial cell to yeast spheroplast cell fusion. As a proof of concept, we demonstrated that the method can be used to capture a piece of genome, or to clone and edit the whole genome from six different Mycoplasma species. This method was then adapted to Bsu, showing for the first-time yeast spheroplast and Gram+ protoplast cell fusion.  A fragment of ~150 kb has been successfully cloned in yeast.

Even if, the entire Bsu genome has not yet been cloned in yeast, several critical elements have been identified. First of all, this work underlines the importance of the cloning method to be adopted depending on the organism of interest. Then, it emphasizes the existence of both biological and technical factors that explain current difficulties and that will have to be taken into account in subsequent experiments. Finally, it enabled the development of the new in-yeast cloning method called CReasPy-Fusion which expands the catalog of technics already described. Through its versatility, it opens up prospects for the capture of large genome fragments, the suppression of problematic loci, and to support the assembly of synthetic fragments.

Jury :

  • M. JULES Matthieu
    Professeur à INRAE, AgroParisTech et Université Paris-Saclay - Président
  • M. RODRIGUE Sébastien
    Professeur agrégé à l’Université de Sherbrooke (QC Canada) - Rapporteur
  • Mme DABOUSSI Fayza
    Directrice de recherche à INRAE-INSA de Toulouse - Rapportrice
  • M. LABROUSSAA Fabien
    Docteur à l’Université de Bern (Suisse) - Examinateur
  • Mme LARTIGUE Carole
    Chargée de Recherche à INRAE de Bordeaux - Directrice de thèse
  • M. BLANCHARD Alain
    Professeur à l’Université de Bordeaux - Directeur de thèse