Allelic diversity in cellulose and lignin biosynthetic genes of Eucalyptus urophylla S.T. Blake

Abstract

Eucalyptus urophylla is one of the most extensively used forest tree species in plantation forestry worldwide. Commonly, E. urophylla is used in hybrid combinations with species possessing better wood properties largely because it is an exceptional grower and it imparts good disease resistance. E. urophylla is endemic to islands of the Lesser Sunda archipelago situated north of Australia. Human induced deforestation practices including urbanization are threatening the existence of several natural populations of the species throughout its range. It has become crucial that efforts be made to conserve the genetic resources in this species. To this end, a forest tree conservation genetics organization called Camcore (http://www.camcore.org) in collaboration with other forestry institutions has initiated seed collection explorations throughout the Lesser Sunda archipelago. Collected seed was sown in provenance test trials to gather information including growth performance of different genotypes in exotic locations. Comprehensive species-wide genetic diversity surveys (at the gene and genome levels) will assist in determining the genetic relationships between different E. urophylla populations, information that is relevant for guiding in situ and ex situ conservation strategies for the species. Nucleotide diversity studies exploit the diversity between homologous gene sequences from different individuals to identify the genetic variation underlying phenotypic traits. Commonly, genetic variation is in the form of single nucleotide polymorphisms (SNPs). Information on SNP diversity coupled with a detailed understanding of the molecular evolution of candidate genes including linkage disequilibrium (LD), selection and recombination may lead to the identification of haplotypes (a combination of SNPs that are inherited together) that associate with trait variation. Thus, sequence diversity surveys in candidate wood biosynthetic genes in E. urophylla may lead to the identification of allelic (SNP) haplotypes that associate with wood quality traits. Such haplotypes will be very valuable in Eucalyptus breeding programmes. The aim of the current M.Sc. study was to investigate levels of nucleotide and allelic (SNP) diversity in three candidate wood biosynthetic genes of E. urophylla. Levels of nucleotide diversity were surveyed in two cellulose biosynthetic genes, namely, cellulose synthase 1 (CesA1) and sucrose synthase 1 (SuSy1) , and the lignin biosynthetic gene cinnamyl alcohol dehydrogenase 2 (CAD2) of E. urophylla. This was achieved by sequencing two DNA fragments of approximately 1000 base pairs (bp) from the 5’ and 3’ ends of one randomly cloned allele (for each gene) in each of the 25 E. urophylla representative individuals. These individuals originated from different families and populations across the seven islands of the Lesser Sunda archipelago. Average levels of nucleotide diversity () and SNP haplotype diversity in EuCesA1, EuSuSy1 and EuCAD2 genes were approximately 1% and 0.95, respectively. SNP density was similar among the three genes with one SNP occurring every 40 bp on average. LD declined to minimal levels within 1000 bp in EuCesA1 and EuSuSy1, but remained significant across the 3000 bp length of EuCAD2. An allele-based geographic analysis based on SNP haplotypes revealed that there was no significant clustering of SNP haplotypes based on island of origin. Nonetheless, high SNP density and low LD levels suggest that the E. urophylla may be useful for high-resolution LD mapping and gene-based marker development for marker-assisted breeding programmes.