**5.4 Identification and characterization of trans-acyltransferse polyketides**

This class of small molecule polyketides is biosynthesized through activity of a freestanding acyltransferases and constitutes one of the most important groups of pharmacologically interesting polyketides. Considering their pharmaceutical

**73**

pathways [108].

*Harnessing the Genetic Diversity and Metabolic Potential of Extremophilic Microorganisms…*

implication and rather limited catalogue from the cultivated microorganism, the metagenomic route of discovery has been adapted. In this approach, the metagenomes from various environments including the extremophilic environments have been probed for presence of a conserved trans-Acyltransferase specific DNA sequences [104]. Using this approach, a single amplicons have been identified which would produce the novel Trans-acyltransferse polyketides. Unlike the Type II polyketides, studies with heterologous transformation and expression of the Trans-acyltransferse polyketides are relative obscure, yet, a few discreet studies have shown genesis of hybrid Trans-acyltransferse polyketides. In one such examples study, a gene encoding for O-methyltransferase from the pederin gene clusters was transformed in a mycalamide-A producing strains. Upon expression the O-methyltransferase catalyzed a site-specifically methylation which resulted in production of a hybrid compound 18-O-methylmycalamide which showed

**6. Single Cell Genome Analyses of the extremophilic microbial diversity**

The single amplified genomes (SAGs) are subsequently used for screening by PCR amplification and NGS sequencing. The taxonomic identity of the concerned extremophilic microbial cell is ascertained with 16S rRNA gene sequencing, whereas subsequent shotgun or NGS sequencing, assembly and annotation is carried out with single amplified genomes of interest identified through preliminary

Despite its tremendous scientific capabilities, the SCGA is yet to make outreaching impact on microbial genomics in general and extremophilic microbiology in particular. The technical procedure used for SCGA faces many challenges that are not yet completely addressed. The most critical challenges include: (i) technical limitation in precise and reproducible separation of single bacterial cells with available methodologies; (ii) low amounts of starting DNA recoverable from single bacterial cell; (iii) requirement of a high degree of amplification; (iv) possibility of cross contamination; (v) introduction of chimeric artifacts and biases in genomic coverage during single genome amplification; and (v) poor post-sequencing quality control, data analyses and sequence assembly [110]. Due to these limitations, the resulting composite assemblies from SCGA can often represent incomplete or inaccurately characterized genomes for a given strain or species [107, 111]. However, several technological updates are being made to circumvent these limitations of the SCGA, which would soon enable highly accurate data generation and its physiological interpretation based on the absence as well as presence of genes and

A recent concept in the field of the culture- independent approaches for identification and characterization of microbial genetic and metabolic diversity is "Single Cell Genome Analyses (SCGA)" [106]. This approach accesses genomes from one cell at a time. Therefore, this approach allows the analyses of the microbial genetic and metabolic diversity at the level of the most fundamental biological unit. The central technical aspect of this approach involves separation of individual cells from a complex mixture of environmental matrix using a cell sorting methods such as fluorescence-activated cell sorting (FACS). Cell separation is followed by cell lysis and recovery of the femtogram levels of DNA from a Single cell. The recovered single cell DNA is amplified using multiple displacement amplification (MDA) and amplification of single cell genomic DNA, such that the quantities of DNA increases


fold increase) [107, 108].

*DOI: http://dx.doi.org/10.5772/intechopen.82639*

significantly improved antitumor activities [105].

to 100s of nano grams – 10s of micro grams (a 103

phylotype characterization [106–109].

*Harnessing the Genetic Diversity and Metabolic Potential of Extremophilic Microorganisms… DOI: http://dx.doi.org/10.5772/intechopen.82639*

implication and rather limited catalogue from the cultivated microorganism, the metagenomic route of discovery has been adapted. In this approach, the metagenomes from various environments including the extremophilic environments have been probed for presence of a conserved trans-Acyltransferase specific DNA sequences [104]. Using this approach, a single amplicons have been identified which would produce the novel Trans-acyltransferse polyketides. Unlike the Type II polyketides, studies with heterologous transformation and expression of the Trans-acyltransferse polyketides are relative obscure, yet, a few discreet studies have shown genesis of hybrid Trans-acyltransferse polyketides. In one such examples study, a gene encoding for O-methyltransferase from the pederin gene clusters was transformed in a mycalamide-A producing strains. Upon expression the O-methyltransferase catalyzed a site-specifically methylation which resulted in production of a hybrid compound 18-O-methylmycalamide which showed significantly improved antitumor activities [105].
