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Dam and Dcm methylations prevent gene transfer into Clostridium pasteurianum NRRL B-598: development of methods for electrotransformation, conjugation, and sonoporation

Overview of attention for article published in Biotechnology for Biofuels, January 2016
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Title
Dam and Dcm methylations prevent gene transfer into Clostridium pasteurianum NRRL B-598: development of methods for electrotransformation, conjugation, and sonoporation
Published in
Biotechnology for Biofuels, January 2016
DOI 10.1186/s13068-016-0436-y
Pubmed ID
Authors

Jan Kolek, Karel Sedlar, Ivo Provaznik, Petra Patakova

Abstract

Butanol is currently one of the most discussed biofuels. Its use provides many benefits in comparison to bio-ethanol, but the price of its fermentative production is still high. Genetic improvements could help solve many problems associated with butanol production during ABE fermentation, such as its toxicity, low concentration achievable in the cultivation medium, the need for a relatively expensive substrate, and many more. Clostridium pasteurianum NRRL B-598 is non-type strain producing butanol, acetone, and a negligible amount of ethanol. Its main benefits are high oxygen tolerance, utilization of a wide range of carbon and nitrogen sources, and the availability of its whole genome sequence. However, there is no established method for the transfer of foreign DNA into this strain; this is the next step necessary for progress in its use for butanol production. We have described functional protocols for conjugation and transformation of the bio-butanol producer C. pasteurianum NRRL B-598 by foreign plasmid DNA. We show that the use of unmethylated plasmid DNA is necessary for efficient transformation or successful conjugation. Genes encoding DNA methylation and those for restriction-modification systems and antibiotic resistance were searched for in the whole genome sequence and their homologies with other clostridial bacteria were determined. Furthermore, activity of described novel type I restriction system was proved experimentally. The described electrotransformation protocol achieved an efficiency 1.2 × 10(2) cfu/μg DNA after step-by-step optimization and an efficiency of 1.6 × 10(2) cfu/μg DNA was achieved by the sonoporation technique using a standard laboratory ultrasound bath. The highest transformation efficiency was achieved using a combination of these approaches; sono/electroporation led to an increase in transformation efficiency, to 5.3 × 10(2) cfu/μg DNA. Both Dam and Dcm methylations are detrimental for transformation of C. pasteurianum NRRL B-598. Methods for conjugation, electroporation, sonoporation, and a combined method for sono/electroporation were established for this strain. The methods described could be used for genetic improvement of this strain, which is suitable for bio-butanol production.

Twitter Demographics

The data shown below were collected from the profile of 1 tweeter who shared this research output. Click here to find out more about how the information was compiled.

Mendeley readers

The data shown below were compiled from readership statistics for 74 Mendeley readers of this research output. Click here to see the associated Mendeley record.

Geographical breakdown

Country Count As %
New Zealand 1 1%
Czechia 1 1%
Unknown 72 97%

Demographic breakdown

Readers by professional status Count As %
Researcher 19 26%
Student > Ph. D. Student 19 26%
Student > Bachelor 10 14%
Student > Master 10 14%
Other 5 7%
Other 2 3%
Unknown 9 12%
Readers by discipline Count As %
Biochemistry, Genetics and Molecular Biology 31 42%
Agricultural and Biological Sciences 21 28%
Veterinary Science and Veterinary Medicine 3 4%
Pharmacology, Toxicology and Pharmaceutical Science 2 3%
Computer Science 1 1%
Other 4 5%
Unknown 12 16%

Attention Score in Context

This research output has an Altmetric Attention Score of 1. This is our high-level measure of the quality and quantity of online attention that it has received. This Attention Score, as well as the ranking and number of research outputs shown below, was calculated when the research output was last mentioned on 23 January 2016.
All research outputs
#5,244,192
of 7,012,411 outputs
Outputs from Biotechnology for Biofuels
#402
of 553 outputs
Outputs of similar age
#218,221
of 317,437 outputs
Outputs of similar age from Biotechnology for Biofuels
#50
of 95 outputs
Altmetric has tracked 7,012,411 research outputs across all sources so far. This one is in the 14th percentile – i.e., 14% of other outputs scored the same or lower than it.
So far Altmetric has tracked 553 research outputs from this source. They receive a mean Attention Score of 3.8. This one is in the 17th percentile – i.e., 17% of its peers scored the same or lower than it.
Older research outputs will score higher simply because they've had more time to accumulate mentions. To account for age we can compare this Altmetric Attention Score to the 317,437 tracked outputs that were published within six weeks on either side of this one in any source. This one is in the 17th percentile – i.e., 17% of its contemporaries scored the same or lower than it.
We're also able to compare this research output to 95 others from the same source and published within six weeks on either side of this one. This one is in the 37th percentile – i.e., 37% of its contemporaries scored the same or lower than it.