Developing markers for Sigatoka leaf spot disease (Mycosphaerella musicola Leach) resistance in banana (Musa spp.)

Abstract

Sigatoka leaf spot (Mycosphaerella musicola Leach) disease is a limiting factor in banana production in India and other places. Breeding for resistance is the most effective method to control Musa diseases. However, Musa improvement using conventional methods has been hampered due to lack of genetic variability, resulting to biotechnological approaches. In this regard, marker-assisted selection has become a reliable method to improve disease resistance in Musa. The objective of this study was to identify markers that may be linked to Sigatoka leaf spot disease in Musa, using RAPDs and converting such into sequence characterized amplified region (SCAR). Consequently, a total of 102 oligonucleotide OPERON primer pairs were used to screen genomic DNA from two resistant cultivars: Calcutta 4 (Musa acuminate, AA) and Manoranjitham (AAA), and two susceptible cultivars Anaikomban (AA) and Grande Naine (AAA) with only 11 (10.8%) of the primers being polymorphic. Eventually, OPK 01 and OPK 11 primers in Calcutta 4 were eluted, but only OPK 11 was sequenced and cloned using pGEM-2T vector, resulting to a band size of 4.3 KB, and the development of two SCAR markers. A FASTA search in the Musa genome database could not identify corresponding gene sequences that show homology with the sequenced PCR fragment. Finally, the SCAR marker was used to amplify genomic DNA from the segregating population which could not discriminate between resistant and susceptible samples. This may be due to amplification conditions, limited number of primers and most importantly, the absence of tight linkage with the gene of interest. In conclusion, it may be necessary to screen the segregating population with more reliable and reproducible amplified fragment length polymorphism.

Key words: Marker-assisted selection, disease resistance, Musa, random amplification of polymorphic DNA (RAPD), genetic improvement, SCAR.