Study On Chloroplast Genome Structure Variation And Screening Of Hypervariable Region In Magnoliaceae

Chloroplast genomes are important components of plant genomes,its conservativity and slow rate of nucleotide substitution make cpDNA a suitable and widely used tool in plant phylogeny and species identification.Most Magnoliaceae plants have excellent horticultural value.The study of chloroplast genome is conducive to interspecific relationships and species identification research in Magnoliaceae.In this paper,the structural variation of chloroplast genome was studied by comparing the composition,structure and sequence similarity in 21 Magnoliaceae plants.On this basis,the distribution of sequence polymorphism at different classification levels was discussed by comparing a large number of microstructural changes among genera,subgenera and species.At the same time,in view of the limited identification ability of international general barcode in Magnoliaceae,this paper makes a preliminary exploration on how to screen hypervariable region gene fragments that meet the requirements of DNA barcode,and provides a reference for the follow-up study of Magnoliaceae DNA barcode.The main research contents and results are as follows:(1)The chloroplast genome of Magnoliaceae exhibited a typical double-stranded structure with four-part regions.The length of Magnoliaceae chloroplast genome ranges from 159,429 bp to160,183 bp,encoding a total of 112 genes,of which 79 protein-coding genes,29 tRNAs and 4 rRNAs.The gene content and gene order were consistent across the chloroplast genomes with no gene rearrangement.No structural variation over 1 kb was found by sequence similarity comparison.(2)The microstructural variation(sequence polymorphism)less than 1 kb in the coding region is mainly in the form of single base mutation,insertion/deletion and simple repeat sequence.Single base mutation was mainly the exchange between G-A/A-G and C-T/T-C,and the two forms accounted for61.5% of the total number of mutations.The amount of insertion/deletion in the coding region had a tendency to increase as the classification level increases,and deletion within the genus level was more likely to occur than insertion.The simple repeat sequence that occursed in the coding region had three independent evolution events.(3)Non-coding region(including intron region)counts the sequence polymorphism of simple repeat sequence,inversion repeat and insertion/deletion which are less than 1 kb.Single nucleotide repeats accounted for 71.7% of simple repeats in non-coding regions(including intron regions),and the length of insertion/deletion for non-coding regions(including intron regions)was mainly within 10 bp.Large segment of insertions/deletions had a tendency to occur at higher classification levels.(4)Mega 6.0 was used to calculate the variation sites and information sites of all coding regions,non-coding regions in Magnoliaceae.We found that the small variation of gene regions could not provide valuable high-variable gene fragments.Eight high-resolution gene fragments were screened out by using mutation site rate,information site rate and comparison length in the non-coding regions,including accD-psaI,rps15-ycf1,psaJ-rpl33,psaI-ycf4,trnL-trnF,trnP-psaJ,trnS-psbZ and trnS-trnG.These hypervariable region gene fragments can be used as key reference fragments for subsequent Genus-specific DNA barcode studies.