A Primer and Guidelines for Shotgun Proteomic Analysis in Non-model Organisms.

During the last decade, we have witnessed outstanding advances in proteomics led mostly by great technological improvements in mass spectrometry field allowing high-throughput production of high-quality data used for massive protein identification and quantification. From a practical viewpoint, these advances have been mainly exploited in research projects involving model organisms with abundant genomic and proteomic information available in public databases. However, there is a growing number of organisms of high interest in different disciplines, such as ecological, biotechnological, and evolutionary research, yet poorly represented in these databases. Important advances in massive parallel sequencing technology and easy accessibility of this technology to many research laboratories have made nowadays possible to produce customized genomic and proteomic databases of any organism. Along this line, the use of proteogenomic approaches by combining in the same analysis the data obtained from different omic levels has emerged as a very useful and powerful strategy to run shotgun proteomic experiments specially focused on non-model organisms. In this chapter, we provide detailed procedures to undertake shotgun quantitative proteomic experiments following either a label-free or an isobaric labeling approach in non-model organisms, emphasizing also a few key aspects related to experimental design and data analysis.