Chromosome-Scale Assembly and Annotation of the Macadamia Genome (Macadamia integrifolia HAES 741)

Abstract

Macadamia integrifolia is a representative of the large basal eudicot family Proteaceae and the main progenitor species of the Australian native nut crop macadamia. Since its commercialisation in Hawaii fewer than 100 years ago, global production has expanded rapidly. However, genomic resources are limited in comparison to other horticultural crops. The first draft assembly of M. integrifolia had good coverage of the functional gene space but its high fragmentation has restricted its use in comparative genomics and association studies. Here we have generated an improved assembly of cultivar HAES 741 (4,094 scaffolds, 745 Mb, N50 413 kb) using a combination of Illumina paired and PacBio long read sequences. Scaffolds were anchored to 14 pseudo-chromosomes using seven genetic linkage maps. This assembly has improved contiguity and coverage, with >120 Gb of additional sequence. Following annotation, 34,274 protein-coding genes were predicted, representing 90% of the expected gene content. Our results indicate that the macadamia genome is repetitive and heterozygous. The total repeat content was 55% and genome-wide heterozygosity, estimated by read mapping, was 0.98% or an average of one SNP per 102 bp. This is the first chromosome-scale genome assembly for macadamia and the Proteaceae. It is expected to be a valuable resource for breeding, gene discovery, conservation and evolutionary genomics.

Comparison of long-read methods for sequencing and assembly of a plant genome

“…Assembly statistics were computed using QUAST v5.0.2 (QUAST, RRID:SCR_001228 ) [ 47 ] with a minimum contig length of 10 kb and the parameters --fragmented --large. The publicly available reference genome of M. integrifolia v2 (Genbank accession: GCA_900631585.1 ) [ 8 ] was used as the reference genome for QUAST. To estimate the base accuracy, QUAST was used to compute the number of mismatches and indels as compared to the Illumina short-read assembly generated by SPAdes.…”

“…The macadamia genus contains 4 species: Macadamia integrifolia, Macadamia tetraphylla, Macadamia ternifolia , and Macadamia jansenii . Macadamia cultivars are diploid (2n = 28), with k -mer–based genome size estimates ranging from 758 Mb for M. tetraphylla [ 7 ] to 896 Mb for M. integrifolia [ 8 ]. The first draft genome assembly of the widely grown M. integrifolia cultivar HAES 741 was constructed from short-read Illumina sequence data and was highly fragmented (518 Mb, 193,493 scaffolds, N50 = 4,745 bp) [ 9 ].…”

“…The first draft genome assembly of the widely grown M. integrifolia cultivar HAES 741 was constructed from short-read Illumina sequence data and was highly fragmented (518 Mb, 193,493 scaffolds, N50 = 4,745 bp) [ 9 ]. An improved HAES 741 assembly was generated using a combination of long-read Pacific Biosciences (PacBio) and paired-end Illumina sequence data (745 Mb, 4,094 scaffolds, N50 = 413 kb) [ 8 ]. The genome assembly of M. tetraphylla was also recently produced using a combination of long-read Oxford Nanopore Technologies (ONT) and short-read Illumina sequence data (751 Mb, 4,335 contigs, N50 = 1.18 Mb) [ 7 ].…”

Improvements in the Sequencing and Assembly of Plant Genomes

“…These assemblies represent significant advances over the highly fragmented genomes previously reported for these species [11][12][13][14]. Advances in long read sequencing using different platforms provide improving options for plant genome sequencing and assembly [15].…”

“…Influence of data volume on assembly for Macadamia species N50 of contigs is plotted against the genome coverage. Genome sizes used to calculate coverage were; M.integrifolia, 895Mb[12]; M janseni, 780Mb[2]; M. tetraphylla, 758Mb[23] and M. ternifolia, 758Mb (not known but assumed the same as M. tetraphylla due to similar assembly size). Influence of data volume on assembly for diverse species.…”