**5. Amplicon sequencing and bioprospecting of metagenomes**

In the amplicon sequencing study, first, soil DNA is extracted, and next, 16S/18S rDNA sequences are amplified using a specific set of particular primers targeting variable areas of 16S/18S rDNA, accompanied by filtration of fragments using magnetic beads [54, 90–92]. Consequently, adapters are ligated, and the library of fragments (clones) is amplified along with the samples are sequenced utilizing NGS platform (**Figure 1**). The dataset obtained after sequencing is used for the identification of microbial diversity [54, 55]. Using NGS and the related software, it is doable to solve extremely complicated microbiota compositions with greater precision and to relate the microbial ecosystem of the soil [16, 55, 92], although, it must be considered for accurate data and analysis interpretation while choosing amplicon sequencing working with marker genes [92].

From the start of metagenomics, the study of novel metabolite/biomolecule (DNA polymerases, cellulases, lipases/esterases, chitinases and antibiotics) from the microbial assembly was its first application, and this has advanced with the development of NGS techniques for calculating comparison between community metagenome, meta-proteomics, and meta-transcriptomics [93, 94]. Techniques for recovering novel metabolite that comprise cloning of the microbial DNA from the environment then constructing a small/large-insert libraries, which can be done either by function-based or sequence-based screening of metagenomic libraries are shown in **Table 1**. The resulting metagenomic libraries subsequently transformed in several hosts like Escherichia coli (mostly), *Sulfolobus solfataricus*, *Thermus thermophilus*, *Streptomyces*, and *Proteobacteria* show significant differences in expression modes [108–111].
