6.11.9 Gene Knockout
This genetic tool is used for gene deletion from the organism. Gene knockouts is
done using insertion approach so that functional capacity of mutant gene is enhanced
using suicidal plasmids, site directed FLP recombines, PCR based approaches, and
transposons so that intentionally specific genes can be inactivated. Literature review
gives some genes that were targeted for the desired phenotype, i.e. aldo-keto
reductase, oxidoreductase, cytochrome related genes, hydroxylamine reductase,
NaOH dehydrogenase, alcohol dehydrogenase, and pyruvate dehydrogenase
(Zhang et al. 2013).
6.11.10 Genomics
This technique is a promising approach for strains improvement. Z. mobilis genome
was sequenced in 2005. This strain has 2,056,416-bp chromosome that is circular
and also contains extrachromosomal material called plasmids. In 2005 other strains
of Z. mobilis complete genome sequence were also reported. Genomics approach is a
great help in targeting a particular gene for a specific purpose (Widiastuti et al.
2011).
6.11.11 Transcriptomic
Transcriptomic study of Z. mobilis is a better way to understand gene function.
Different techniques like DNA array or DNA sequencing approaches have been used
in differential gene expression under different stress conditions. Transcriptomic
profiling done in the past was of greater help in the gene regulation. Different studies
were done using ZM4 growing in ethanol stress media, heat shocked media, high
ethanol concentration, and rich and minimal media. Different mutants were made
using genomic profiling like ACR that is sodium acetate tolerant mutant.
6.11.12 Using Shuttle Vectors
A gene important for the production of different alcohols especially isobutanol is
cloned into shuttle vector PEZ15A sp. By incorporating tetracycline inducible
promoter sequence and then introducing it in Z. mobilis, a recombined strain
ZM4-Kit can be formed. This can be confirmed by gene Sanger Sequencing.
Different experiments were performed to check the impact of kdcA gene expression
due to pet promoter for isobutanol production and it was recorded that isobutanol
6 Engineering of Zymomonas mobilis for Enhanced Biofuel Production
175
This genetic tool is used for gene deletion from the organism. Gene knockouts is
done using insertion approach so that functional capacity of mutant gene is enhanced
using suicidal plasmids, site directed FLP recombines, PCR based approaches, and
transposons so that intentionally specific genes can be inactivated. Literature review
gives some genes that were targeted for the desired phenotype, i.e. aldo-keto
reductase, oxidoreductase, cytochrome related genes, hydroxylamine reductase,
NaOH dehydrogenase, alcohol dehydrogenase, and pyruvate dehydrogenase
(Zhang et al. 2013).
6.11.10 Genomics
This technique is a promising approach for strains improvement. Z. mobilis genome
was sequenced in 2005. This strain has 2,056,416-bp chromosome that is circular
and also contains extrachromosomal material called plasmids. In 2005 other strains
of Z. mobilis complete genome sequence were also reported. Genomics approach is a
great help in targeting a particular gene for a specific purpose (Widiastuti et al.
2011).
6.11.11 Transcriptomic
Transcriptomic study of Z. mobilis is a better way to understand gene function.
Different techniques like DNA array or DNA sequencing approaches have been used
in differential gene expression under different stress conditions. Transcriptomic
profiling done in the past was of greater help in the gene regulation. Different studies
were done using ZM4 growing in ethanol stress media, heat shocked media, high
ethanol concentration, and rich and minimal media. Different mutants were made
using genomic profiling like ACR that is sodium acetate tolerant mutant.
6.11.12 Using Shuttle Vectors
A gene important for the production of different alcohols especially isobutanol is
cloned into shuttle vector PEZ15A sp. By incorporating tetracycline inducible
promoter sequence and then introducing it in Z. mobilis, a recombined strain
ZM4-Kit can be formed. This can be confirmed by gene Sanger Sequencing.
Different experiments were performed to check the impact of kdcA gene expression
due to pet promoter for isobutanol production and it was recorded that isobutanol
6 Engineering of Zymomonas mobilis for Enhanced Biofuel Production
175