Genetic systems development in the clostridia

Abstract

This review describes recent developments in the genetic manipulation of the solventogenic clostridia, Clostridium acetobutylicum and C. beijerinckii. It is to be noted that our laboratory stock of C. acetobutylicum ATCC 824, which was obtained from the American Type Culture Collection, has recently been re-identified as C. beijerinckii NCIMB 8052 based on DNA similarity studies using the S1 nuclease method (personal communication, Dr. Jiann-Shin Chen, Virginia Polytechnic Institute and State University). Reference to our laboratory 824 culture has been changed to C. beijerinckii NCIMB 8052 throughout this paper in order to be consistent with this finding. The focus of this review specifically involves the characterization of an M13-1ike genetic system for the clostridia based on the pCAK1 phagemid, as well as preliminary work on development of a plasmid-based vector based on the indigenous pDM11 plasmid recovered from C. acetobutylicum NCIB 6443. The construction of a C. beijerinckii strain with amplified endoglucanase activity was achieved by inserting the engB gene from C. cellulovorans into C. beijerinckii. The successful expression of a heterologous engB gene from C. cellulovorans in C. beijerinckii NCIMB 8052 has important industrial significance for the eventual utilization of cellulose by this acetone-butanol-ethanol fermentation microorganism.

Biobutanol: science, engineering, and economics

“…Blaschek and White [32], and Girbal and Soucaille [33]. Park and Geng [34] and Belafi-Bako et al [35] have reviewed the latest developments with respect to various techniques (vaccumization, gas stripping, liquid-liquid extraction, and adsorption) used for simultaneous product removal and recovery from fermentation broth.…”

“…This review will touch upon various aspects of ABE fermentation, including the following: (i) microbial cultures for fermentation and metabolic pathway; (ii) substrates for fermentation; (iii) fermentation protocols and reactor design (including immobilized systems); (iv) solvent recovery techniques; (v) mathematical modeling; and (vi) economics of the process. Genetic and metabolic engineering aspects of clostridial species is outside the scope of this review, and reader is referred to the reviews by Blaschek and White [32], Woods [31], Ezeji et al [51], and Lee et al [11]. Moreover, this review mainly focuses on literature published since 1985; for earlier literature, we refer the reader to Jones and Woods [12].…”

“…Development of an economic and efficient ABE fermentation technology is possible only through concerted efforts of microbiologists, biotechnologists, and chemical engineers. Development of genetically modified strain having higher solvent tolerance and generation of metabolically engineered strains capable of utilizing varied carbon source and producing higher amount of butanol rather than other side products are the challenges set for scientific community . As mentioned in the previous sections, co‐culturing of clostridia and simultaneous saccharification and fermentation are few of the technologies, which on successful implementation can lead to economic acceptance of the process at large scale .…”

“…and McNeil and Kristiansen , and Maddox . Major reviews on genetic engineering and manipulation of the clostridia, cloning, and DNA sequencing published in 1990s include those from Blaschek , Woods , Blaschek and White , and Girbal and Soucaille . Park and Geng and Belafi‐Bako et al .…”