EST Based Gene Expression Analysis In Porphyra Haitanesis Sporophyte And Super-High-Yield Hevea Brasiliensis Latex With Further Exploration The High Yield Mechanism Of Rubber

Expressed sequence tag (EST), a short sub-sequence of a transcribed spliced nucleotide sequence, is produced by one-shot sequencing of an end of cloned mRNA. The length of EST usually ranges from 100 to 500 base pairs which are limited by the current technologies. Due to the low-cost, high speed and high throughout, EST technology has rapidly developed since it firstly reported in 1991, and EST sequence has become the most rapidly increased data in public databases. Nowadays, EST technology has been widely used in molecular genetics, genomics and many other fields, playing an important role in researches such as gene expression profile, identification of new gene, construction of gene map, and detection of single nucleotide polymorphisms and so on. In this study, we profiling the gene expression pattern of Porphyra haitanensis filamentous sporophyte and super-high-yield Hevea brasiliensis latex based on the EST data.Rubber is an important raw material in both industry and stratagem, The super-high-yield Hevea brasiliensis , discovered in rubber plantation of Yunnan China, is invaluable in researching rubber biosynthesis mechanism and identifying candidate genes related with high yield. In this study, 20126 high quality expressed sequence tags (ESTs) were generated from the latex of the super-high-yield Hevea brasiliensis by sequenced from 5'cDNA and these ESTs were further assembled into 6606 tentative consensus sequences TCs, including 3120 contigs and 2986 singlets. Blast search showed that 5263 of the TCs were highly similar to 3075 known or predicted proteins; whereas the rest of 1343 TCs had no significant similarity to any sequence in the database. Most of the high redundant TCs were involved in biosynthesis and response to stress, such as REF(2290 ESTs), Hevein(919 ESTs), SRPP(525 ESTs), HSP(151 ESTs). Function analysis showed that genes involved cellular metabolic process, secondary metabolic process, macromolecule metabolic process, biosynthesis and defense response were highly represented. Thirteen proteins, including REF, SRPP, FPS, GGPS and all enzymes in the MEV pathway, were found in our library. In contrast, only one enzyme in MEP pathway was detected, suggesting that the MEV might be the dominant pathway in rubber biosynthesis in latex vessels of Hevea brasiliensis. Comparison of super-high-yield latex and general latex revealed that 65 genes have evidently differential expression, in which 11 genes were highly expressed and 54 genes lowly expressed in super-high-yield Hevea brasiliensis. The highly expressed genes mainly participates in biosynthesis, response to stimulus, defense response and macromolecule localization. Within all the thirteen proteins that are important in rubber biosynthesis, REF, SRPP, MK, PMK and IPPI were more abundantly present in our database, indicating their potential importance in high yield.Porphyra has a significant value for economic and scientific research. Recently it has been recognized as a model system for fundamental and applied studies in marine biological sciences. This model organism has a typical dimorphic life cycle consisting of two generations, gametophyte and sporophyte, with completely different morphologies. P. haitanensis is a unique species for China. In this study, EST technology was used to investigate the global gene expression profile of P. haitanensis sporophyte and differential express patterns between sporophyte and gametophytic. A total of 11,000 clones from the sporophyte cDNA library of P. haitanensis were sequenced and generated 5344 high quality ESTs, which were clustered into 2182 tentative consensus sequences (TCs). Database searching showed that 690 (32%) of the 2182 TCs have been identified their homologous genes from Genbank NR database (Nr or Nt, pls correct by yourself) , and 1492 (68%) genes were unique for P. haitanensis. Function analysis indicated that the cellular process and metabolic process were two main functional groups of 33% known function genes. Other important functional groups also included binding, catalytic activity, macromolecular complex, cell part, and hydrolase activity. Metabolic pathway analysis showed that enzymes but PEPC involved C4 carbon fixation pathway were all highly represented, especially the PEPCK, which proved the existence of C4 carbon fixation pathway. Though sporophytes of P. haitanensis and P. yezoensis were highly similar in morphology, we still observed significant differences between the two species at molecular level. Only～28% of P. haitanensis genes identified from P. haitanensis showed high similarity to that of P. yezoensis. Four heat shock genes were only found in P. haitanensis, which may relevant to the high temperature tolerance. The codon usage analysis of 328 full-length cDNA sequences suggested that the genome of P. haitanensis has been exposed to high GC pressure during evolution. In total, 3,680 ESTs derived from this study had been registered in Genbank (AC number from DN604790 to DN608469). This is the most extensive collection of P. haitanensis ESTs, and it will be instrumental for other researchers to understand the unique properties of this model organism and its potentially generation change related genes.