Comparison of two PCR-based DNA markers with high resolution melt analysis for the detection of genetic variability in selected quality protein maize inbred lines

Abstract

Molecular markers are fast, efficient and reliable in detecting distinct differences between genotypes at DNA level. In particular, polymerase chain reaction (PCR)-based markers are widely preferred for genotype characterization in diverse crop species, including maize. It is impossible, however, to obtain the entire information required in plant breeding and conservation programmes solely from a single marker system without integration with other technique(s). The current study was, therefore, initiated to compare results from random amplified polymorphic DNA (RAPD) and simple sequence repeats (SSR) markers in combination with high resolution melt analysis for effectiveness in detecting genetic variability or similarity among selected QPM inbred lines. Eight fresh leaf samples (two per line) were collected from each line for DNA extraction. The genomic DNA extracts were subjected to polymerase chain reaction (PCR) and high resolution melt analyses which involved three RAPD primers and three SSR primer pairs using a Real-Time PCR System. The curves of the HRM analysis showed fairly similar melt patterns among the lines for both types of marker systems. Results obtained from the gelelectrophoresis and genetic distance measures for RAPD markers were also in agreement with the HRM analysis. In general, both marker systems in combination with HRM analyses were able to detect genetic variations within and between the inbred lines, thus justifying the potential use of HRM analysis and indicating the possibility of excluding the use of gel-electrophoresis at the early stage of screening of large number of samples of newly developed inbred lines in order to save time and resources.