Sequencing And Comparative Study Of Chloroplast Genome Of Neochloris Oleoabundans With Several Oleaginous Green Microalgae

With the development of economics and rapid increase of the world population,dramatic increase of global consumption of energy and food has caused severe problems including lack of resources,deterioration of ecological environment,continuous reduction of cultivated land,etc.The cultivation of microalgae is less affected by region and season,and does not occupy cultivatable land.Neochloris oleoabundans has high capacity for oil production and potentiality to produce edible oil as new food resources,but researches on the biology of this oleaginous microalga,especially on molecular biology,are still limited.Studies on chloroplast genome benefit taxonomy,genetic engineering,and food application of microalgae.In this paper,the conditions of chloroplast isolation were optimized,the chloroplast genome was extracted and sequenced,and the gene analysis was performed comparing with several oleaginous green microalgae.This research can provide a basis for the understanding of the biology of N.oleoabundans,genetic engineering of oleaginous microalgae for edible oil,and high efficient genetic-expression of bioactives in chloroplast.The main research results are as follows:(1)The percoll density gradient centrifugation method was used to separate the chloroplasts of N.oleoabundans and the separation conditions were optimized as percoll density of 10%,35% and 50% combined with a suitable set of centrifugation speed operation.(2)Sequencing and segment assembling of N.oleoabundans chloroplast genome revealed the total length of the genome to be 129,716 bp,and gene annotation has indicated 45 genes yet.Analysis of simple repeat sequences showed that N.oleoabundans chloroplast genome contained 242 SSRs(simple sequence repeats),among which 222 were single nucleotide SSRs,and 91.7% of them were A/T single base repeats.(3)Codon preference analysis in chloroplast genomes of N.oleoabundans and Chlorella sorokiniana showed that natural selection was the main factor affecting the codon preference of both microalgae,the optimal codons of N.oleoabundans were TTA,ATT,GTA,TCT,CCA,ACT,GCA,AAA,GAA,TGT,CGT and GGT,and the optimal codons of C.sorokiniana were TTT,TTA,ATT,GTT,TCA,CCA,ACA,GCT,TAT,TGA,CAT,CAA,AAT,AAA,GAT,GAA,CGT and GGT.(4)Phylogenetic analysis showed that the evolutionary relationship of N.oleoabundans was closest to Ettlia genus and far from Neochoris genus.