Study On Pigment Comparison And Differentially Expressed Genes Between Red And White Pitaya Fruit

Pitaya is a cactus(Cactaceae), (Hylocereusundatus). According to the flesh color , it is mainly divided into two kinds of pitaya red meat and white meat. Red pitaya contains rich betacyanin and anthocyanins, and which has a good antioxidation action. According to the reports, there differences in flesh color mainly reflected in the expression of metabolic enzymes and the regulation of transcription factors in the developing process of plants. In this study,we selected pitaya fruit as genetic material of the different growth between red and white pitaya fruits, in green ripe stage and mature stage of pitaya sampling. On the one hand, there extracted and determined eight item nutrition index, like ascorbic acid,carotenoid, soluble protein, soluble sugar, total phenolics, flavonoids, betacyanin and anthocyanin in the pulp, compared green ripe stage with mature stage in red and white pulp.On the other hand, by using bioinformatics of a new generation of high-throughput RNA-Seq sequencing technology, we compared the transcriptome of red and white pitaya during fruit developing stage, had screened differentially expressed genes. According to the analysis, we had found the genes controlling pitaya color. And we cloned the genes and analysised their expression quantity, to explore the differences of pitayas color metabolism.The results are as follows:1 . Comparing the compositions of red and white pitaya, both beet red pigment and anthocyanin were obvious difference, other compositions, such as ascorbic acid,carotenoid, soluble protein, soluble sugar, total phenolics and flavonoids were not significantly difference. the results showed that the different compositions between red and white pitaya were mainly the difference of pigment metabolism.2 . Sample W1 (Green ripe stage of white pitaya), W3(Mature period of white pitaya),R1 (Green ripe stage of red pitaya) and R3(Mature period of red pitaya) respectively got high quality reads 57575196 (97.13%), 55265234 (97.31%), 59436460 (97.55%) and 43087310 (97.9%). which can be assembled with high quality reads, respectively 5914708, 13946140, 14364654, 5296884. The total assembly were 53240 Unigenes,the average length was 840.92bp, N50 was 1429bp, the length distribution was between 228-15381bp. And four samples respectively have 35618(66.90%), 45142(84.79%), 41265 (77.51%) and 38588 (72.48%) Unigenes. 27907 Unigenes were annotated in four databases, and Nr, Swiss-Prot, KEGG and KOG, respectively has 27725, 19279, 16942 and 7394 Unigens, and there are 6065 Unigenes annotated in four database.3 . Analysised the Unigenes from assembly, we got 389 species in protein homology analysis of species. From KOG database analysising we got 24 categories, most of them involved in general function prediction of protein. GO enrichment analysis,molecular function mainly enrichment in catalytic activity and binding function, which respectively were 6496 and 6065 Unigenes. TF analysis, there were 5460 Unigenes involved in transcription factor, and the top ten were Bhlh, MYB and NAC types, they were related to transcription factors which involved in plant pigment metabolism.4 . According to GO function Enrichment analysis of differentially expressed genes between red and white in different stages of pitaya plup, it indicated that the mainly significant enriched in the metabolic pathway, oxidative phosphorylation, flavone and flavonol biosynthesisthe metabolic pathway of tyrosine. These paths and beet pigment synthesis were closely related. And known from pathway analysis of them, we found the most significant was the biosynthesis of phenylpropanoids. Studies shows that the synthetic precursor of beet pigment is based on the metabolism of phenylalanine. It indicted the difference of red and white pitaya pulp is mainly the expression of transcription factors.5. Combined with the KEGG database, we selected 18 Unigenes from metabolic pathway of beet pigment and anthocyanin which corresponding to the 18 related enzyme to do qRT-PCR verification. And there draw the possible metabolic pathway for pigment difference of pitaya.6. Through bioinformatics analysis of Unigene0031379 gene sequence, obtained 915bp length open reading frame, and named U31379M2 By cloning U31379M2 gene,sequencing and bioinformatics analysis showed that U31379M2 is close to the beet species, and it found there was high similarity to carbonic anhydrase, showed that the gene may be involved in the regulation of gene expression of beet pigment.