ABSTRACT
Guava (Psidium guajava L.), a rich source of nutrients, is an important tropical and subtropical fruit of the Myrtaceae family and exhibits magnificent diversity. Genetic diversity analysis is the first step toward the identification of parents for hybridization, genetic mapping, and molecular breeding in any crop species. A diversity analysis based on whole-genome functional markers increases the chances of identifying genetic associations with agronomically important traits. Therefore, here, we sequenced the genome of guava cv. Allahabad Safeda on an Illumina platform and generated a draft assembly of ~304 MB. The assembly of the Allahabad Safeda genome constituted >37.95% repeat sequences, gene prediction with RNA-seq data as evidence identified 14,115 genes, and BLAST n/r, Interproscan, PfamScan, BLAST2GO, and KEGG annotated 13,957 genes. A comparative protein transcript analysis of tree species revealed the close relatedness of guava with Eucalyptus. Comparative transcriptomics-based SSR/InDel/SNP-PCR ready genome-wide markers in greenish-yellow skinned and white fleshed-Allahabad Safeda to four contrasting cultivars viz apple-color-skinned and white-fleshed-Lalima, greenish-yellow-skinned and pink-fleshed-Punjab Pink, purple-black-skinned and purple-fleshed-Purple Local and widely used rootstock-Lucknow-49 were developed. The molecular markers developed here revealed a high level of individual heterozygosity within genotypes in 22 phenotypically diverse guava cultivars. Principal coordinate, STRUCTURE clustering, and neighbor-joining-based genetic diversity analysis identified distinct clusters associated with fruit skin and flesh color. The genome sequencing of guava, functional annotation, comparative transcriptomics-based genome-wide markers, and genetic diversity analysis will expand the knowledge of genomes of climacteric fruits, facilitating trait-based molecular breeding and diversifying the nutritional basket.
ABSTRACT
BACKGROUND: Guava (Psidium guajava L.) is an important fruit crop of tropical and subtropical areas of the world. Genomics resources in guava are scanty. RNA-Seq based tissue specific expressed genomic information, de novo transcriptome assembly, functional annotation and differential expression among contrasting genotypes has a potential to set the stage for the functional genomics for traits of commerce like colored flesh and apple color peel. RESULTS: Development of fruit from flower involves orchestration of myriad molecular switches. We did comparative transcriptome sequencing on leaf, flower and fruit tissues of cv. Allahabad Safeda to understand important genes and pathways controlling fruit development. Tissue specific RNA sequencing and de novo transcriptome assembly using Trinity pipeline provided us the first reference transcriptome for guava consisting of 84,206 genes comprising 279,792 total transcripts with a N50 of 3603 bp. Blast2GO assigned annotation to 116,629 transcripts and PFam based HMM profile annotated 140,061 transcripts with protein domains. Differential expression with EdgeR identified 3033 genes in Allahabad Safeda tissues. Mapping the differentially expressed transcripts over molecular pathways indicate significant Ethylene and Abscisic acid hormonal changes and secondary metabolites, carbohydrate metabolism and fruit softening related gene transcripts during fruit development, maturation and ripening. Differential expression analysis among colored tissue comparisons in 3 cultivars Allahabad Safeda, Punjab Pink and Apple Color identified 68 candidate genes that might be controlling color development in guava fruit. Comparisons of red vs green peel in Apple Color, white pulp vs red pulp in Punjab Pink and fruit maturation vs ripening in non-colored Allahabad Safeda indicates up-regulation of ethylene biosynthesis accompanied to secondary metabolism like phenylpropanoid and monolignol pathways. CONCLUSIONS: Benchmarking Universal Single-Copy Orthologs analysis of de novo transcriptome of guava with eudicots identified 93.7% complete BUSCO genes. In silico differential gene expression among tissue types of Allahabad Safeda and validation of candidate genes with qRT-PCR in contrasting color genotypes promises the utility of this first guava transcriptome for its potential of tapping the genetic elements from germplasm collections for enhancing fruit traits.
