Organelle Genome Assembly,Analysis,and DNA Barcoding Development Of Carya Illinoinensis

The chloroplast genome and mitochondrial genome,also known as organelle genomes,are important components of the plant genome.As the control centers of plant chloroplasts and mitochondria,organelle genomes carry the important mission and influence of the physiological and biochemical responses of plant cells.The assembly and analysis of the chloroplast and mitochondrial genomes of pecan(Carya illinoinensis)will help gain a deeper understanding of this species and protect its germplasm identification.The abundant variety resources and indistinguishable morphological characters of pecan have posed obstacles to the management and breeding of this species.Thus,the development of DNA barcodes for species identification using organelle genomes may be helpful.In this study,we had assembled the chloroplast and mitochondrial genome of pecan and analyzed their genome features.The results of organelle genome analysis could provide a theoretical reference for the further study of pecan.We then developed and designed DNA barcoding groups that can identify multiple pecan cultivars based on the variations in the organelle genome.The main results of this study are shown as follows:(1)The chloroplast genomes of seven pecan cultivars(Pawnee,Mahan,Mohawk,Schley,Starking,Stuart,Success)were successfully assembled and analyzed for comparison.The chloroplast genome of pecan was 160 819 bp in length and 36.1% in GC content.It encodes a total of 130 genes,containing 90 SSR repeat sites,and has a high preference for amino acids such as serine and leucine.The other cultivars remained highly consistent,and no differential information was detected except the chloroplast genome of Starking cultivar with a missing 300 bp in length.(2)The mitochondrial genomes of Pawnee cultivar were successfully assembled using Pacbio data.The total length of the mitochondrial genome of pecan was 495 514 bp,with a GC content of 45%.It encodes 73 genes,including 166 SSR repeat sites with a preference for amino acids such as leucine and valine,lacking the tRNA gene for leucine transport.(3)A collinearity analysis was performed between the organelle and nuclear genome.It was found that there were long migratory sequences between the chloroplast genome and mitochondrial genome of pecan.Both two organelle genomes had common sequences with the nuclear genome,and both had a different preference for chromosomes.However,we found no gene exchange in those migratory sequences.(4)A total of 34 pairs of amplifiable DNA barcodes were developed based on the organelle genome and ITS2 sequences of pecan.The gel electrophoresis experiments showed that the ITS2 sequence barcodes could distinguish multiple cultivars into two taxa.After analyzing the variant loci in the sequencing results,we found that the barcode group of ITS2 + trnT-psbD + nad6 can be used to distinguish the experimental species.