Protein stable isotope probing with H 2 18 O differentiated cold stress response at permissive temperatures from general growth at optimal conditions in Escherichia coli K12

2020 ◽  
Vol 35 (1) ◽  
Author(s):  
Robert Starke ◽  
Stephanie Serena Schäpe ◽  
Nico Jehmlich ◽  
Martin Bergen
2020 ◽  
Vol 66 (8) ◽  
pp. 491-494
Author(s):  
Severin Weis ◽  
Sylvia Schnell ◽  
Markus Egert

RNA-based stable isotope probing (RNA-SIP) is used in molecular microbial ecology to link the identity of microorganisms in a complex community with the assimilation of a distinct substrate. The technique is highly dependent on a reliable separation of isotopic-labeled RNA from unlabeled RNA by isopycnic density gradient ultracentrifugation. Here we show that 13C-labeled and unlabeled Escherichia coli RNA can be sufficiently separated by isopycnic ultracentrifugation even in the absence of formamide. However, a slightly lower starting density is needed to obtain a distribution pattern similar to that obtained when formamide was used. Hence, the commonly used addition of formamide to the centrifugation solution might not be needed to separate 13C-labeled RNA from unlabeled RNA, but this must be verified for more complex environmental mixtures of RNA. Clearly, an omission of formamide would increase the safety of RNA-SIP analyses.


2011 ◽  
Vol 146 (2) ◽  
pp. 163-169 ◽  
Author(s):  
Sinisa Vidovic ◽  
Anil K. Mangalappalli-Illathu ◽  
Darren R. Korber

2011 ◽  
Vol 1 ◽  
pp. 1060-1066 ◽  
Author(s):  
Iris Cornet ◽  
Eva Van Derlinden ◽  
Astrid M. Cappuyns ◽  
Wim Bruyninckx ◽  
Agnes Kovacs ◽  
...  

2010 ◽  
Vol 76 (7) ◽  
pp. 2203-2211 ◽  
Author(s):  
Ana Maria Moreno ◽  
Carsten Matz ◽  
Staffan Kjelleberg ◽  
Mike Manefield

ABSTRACT It is well understood that protozoa play a major role in controlling bacterial biomass and regulating nutrient cycling in the environment. Little is known, however, about the movement of carbon from specific reduced substrates, through functional groups of bacteria, to particular clades of protozoa. In this study we first identified the active protozoan phylotypes present in activated sludge, via the construction of an rRNA-derived eukaryote clone library. Most of the sequences identified belonged to ciliates of the subclass Peritrichia and amoebae, confirming the dominance of surface-associated protozoa in the activated sludge environment. We then demonstrated that 13C-labeled protozoan RNA can be retrieved from activated sludge amended with 13C-labeled protozoa or 13C-labeled Escherichia coli cells by using an RNA stable isotope probing (RNA-SIP) approach. Finally, we used RNA-SIP to track carbon from bicarbonate and acetate into protozoa under ammonia-oxidizing and denitrifying conditions, respectively. RNA-SIP analysis revealed that the peritrich ciliate Epistylis galea dominated the acquisition of carbon from bacteria with access to CO2 under ammonia-oxidizing conditions, while there was no evidence of specific grazing on acetate consumers under denitrifying conditions.


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