The Search for possible endophytes in the growth zone of maize leaves

Abstract

Zea mays is one of the most important agricultural crop-species for the human diet, but little is known about the endophytes associated with maize. Within the last few years, endophytes are becoming more interesting as they can change our understanding about the relationship between bacteria and plants. There are a lot of studies with regards to this topic from the last couple of years, but not much is known on the role that these microorganisms develop colonising a particular cultivar, or their interaction with the host’s genome. The main objective of this project was to find evidence of bacterial endophyte organisms, and developing a PCR based protocol for isolating bacteria within the leaf growth zone of maize. This area has been chosen for this project since it has not been studied before in terms of endophytes. This project describes non-maize related sequences, extracted from maize leaf material, which could provide evidence for endophytes. In order to find robust evidence, firstly, next generation sequencing (NGS) data were mapped against the maize reference genome [Taxon ID: 4577]. The sequences that did not align against the maize reference genome, were the ones used during the study. These sequences were BLASTed (Boratyn , et al. 2012) using the NCBI database to get a taxonomic report. The most significant alignment was for Desulfitobacterium hafniense. To support this, various PCR protocols were designed by using two different kind of primers: general bacterial primers (A) and specific bacterial primers (B), which avoid chloroplast gene amplification. These were then sequenced with Illumina. The best alignments for general bacterial (A) primers were for 16S rRNA chloroplast genes and uncultured bacteria, which did not give useful information. However, when using the specific bacterial primers (B), the best hits were for uncultured bacteria clone FFCH13347. Since we were not able to get the same result as in the NGS data taxonomic study, we are not able to confirm the presence of Desulfitobacterium hafniense in the leaf growth zone or the presence of other bacteria organisms. Further analysis about possible endophytes within the leaf growth zone are thus needed