Table 2.

Molecular biology techniques and their pros and cons for studying plant-microbe interactions.

NGS Techniques	Softwares	Pros	Cons	Purposes	References
Genomics and Metagenomics	Assembly: Meta Velvet and Ray Meta, IDBA-UD Profiling: AMPHORA, MetaPhlAn mOTU Function Analysis: IMG/M, MG-RAST And CAMERA	Profiling is unbiased; Uncultured microbial studies can be performed; Correlation can be observed between genes of diverse organisms from similar environment	Less information related to sequencing of the marker genes; Less abundance anticipating functions of the gene are not equivalent to protein content expressed	New species along with their taxonomic profile will be discovered; Potential, metabolic, functional and evolutionary relationships will be derived; Helps in genome reconstruction	[80-84]
Transcriptomics and Metatranscriptomics	Mapping: BWA-SW; Bowtie2 de Novo Assembly: IDBA-MT Function Analysis: CAMERA and MG-RAST	Novel transcripts determination and sensitive methods for detection; Aids in detecting simultaneous expression of the host and microbial gene; Simple to perform; Metatranscriptomics allows capturing of transcriptomes in case of un-cultured bacteria	Low concentration of rRNA presence in the samples; Difficult to assign transcripts to specific organisms; High complexity of the community and host transcriptome leads to low sensitivity; Extensive genome is needed for mapping	Analysis of Pathways and active function study
Proteogenomics	Mascot: Protein Identification; MG-RAST and camera	Proper estimation of functional activities in comparison to transcriptomics; Results in Semi-quantification of the proteins in the environment; More accurate to perform qualitative protein analysis	Reference genes needed for identification of proteins; Difficulty in sample preparation process; Quantity of various proteins cannot be compared; Difficult for performing in plants due to host contamination	Determination of functions and analysis of pathways
Metabolomics	Metabolite study	Metabolites produced due to plant-microbe interactions can be identified; Nominal bias analysis of various compounds	Size of reference public databases is limited; Different metabolites with their functions give similar signals during Mass spectrometry; Similarity between primary metabolites of microbes and plants results in their difficult determination
Marker gene	Amplicon Noise, mothur, QIIME	Classification of new and rare species	Problems during amplification in PCR	Novel species with taxonomic profiling can be discovered