OBJECTIVE To study the reasons that the seedless fruits of Siraitia grosvenorii developed to smaller ones. METHODS The differences of fruit expanding, gene expressing and cell development were investigated between the triploid and diploid fruits from two strains F050 and F049. RESULTS The results showed the expanding of triploid fruits was stopped about 20 days after artificial pollination, 10 days earlier than the diploid fruits. Meanwhile, it was also investigated that aux expressing level in the triploid fruits was greatly higher than that in diploid fruits, while its ipt, cyt-p450, spds, cycB, cycD1, cycD3, cdkA, cdkB, exp and xth expressing level were greatly lower than that in diploid fruits. The majority of sarcocarp cells of the triploid fruits kept in the stage of small ones comparing with the diploid fruits. CONCLUSION The expression of the genes of the proteins involving cell division and expansion was inhibited as the significant reduction of endogenous IAA, CTK, GA and SPDS after bad fertilization and the embryo abortion, which resulted in the end of division and extension of sarcocarp cells. Above reasons induce small fruits of triploid seedless Siraitia grosvenorii.

This study analyzed the differences in karyotype and genetic variation between a mutant and wild-type Siraitia grosvenorii. Genetic variation included changes in genome and gene expression by SRAP molecular markers. Results showed that wild-type S. grosvenorii was diploid, with a chromosome number of 2n = 2x = 28, whereas the mutant was tetraploid with a chromosome number of 2n = 4x = 56. 4573 DNA bands were obtained using 189 different primer combinations, 577 of which were polymorphic, averaging 3.1 bands for each primer pair, while 1998 pairs were identical. There were no apparent differences on bands amplified by most primer pairs. After comparing the diploid and tetraploid strains, the data generally indicated that the polymorphism would be quite low. 2917 cDNA bands were generated using 133 primer combinations, and stable and clearly differential fragments were sorted out, cloned and sequenced. Ninety-two differentially expressed fragments were successfully sequenced. Sequence analysis showed that most fragments had significant homologous nucleotide sequences with resistant to stress and photosynthesis genes, including ribulose-1,5-bisphosphate carboxylase/oxygenase, phosphoenolpyruvate carboxykinase, pyruvate kinase, peroxisomal membrane transporter, NBS-LRR type resistance protein, protein phosphatase and others. The results revealed that the tetraploid strain has more resistant and photosynthesis ability than its diploid relatives, which providing reference information and resources for molecular breeding and seedless Luohanguo.

In order to reveal the differentially-expressed genes involved in triterpenoid saponin biosynthesis of Siraitia grosvenorii,50 DAF(days after flowering)and 70 DAF fruits from different developmental stages were employed to construct subtractive cDNA library by suppression subtractive hybridization(SSH)in this study.Total 641 positive clones were selected randomly and sequenced from the forward-subtracted cDNA library(70 DAF as the tester and 50 DAF as the driver),and 622 high quality sequences were obtained.The rate of the recombination fraction was above 96%.The distribution of inserted fragments ranged from 101 bp and 934 bp,and the average fragment size was 500 bp.Followed by BLASTN and BLASTX for sequences dates,201 ESTs showed no significant matches to any other sequences in NCBI nucleotide sequence database,they were probably novel genes.The other 421 ESTs carried with remarkable identity to proteins with known function in the database,which fell into several functional categories including energy and secondary metabolism,transcription factors,ripening,senescence and pathogen-resistance.Further results indicated that the cDNA library quality were conformed to SSH library standard and could be used in further studies,also would provide reference for studying the differentially expressed genes,exploring molecular mechanism of triterpenoid saponin biosynthesis,and increasing productivity and quality of mogrosides in S.grosvenorii.

BACKGROUND:Siraitia grosvenorii (Luohanguo) is an herbaceous perennial plant native to southern China and most prevalent in Guilin city. Its fruit contains a sweet, fleshy, edible pulp that is widely used in traditional Chinese medicine. The major bioactive constituents in the fruit extract are the cucurbitane-type triterpene saponins known as mogrosides. Among them, mogroside V is nearly 300 times sweeter than sucrose. However, little is known about mogrosides biosynthesis in S. grosvenorii, especially the late steps of the pathway.RESULTS:In this study, a cDNA library generated from of equal amount of RNA taken from S. grosvenorii fruit at 50 days after flowering (DAF) and 70 DAF were sequenced using Illumina/Solexa platform. More than 48,755,516 high-quality reads from a cDNA library were generated that was assembled into 43,891 unigenes. De novo assembly and gap-filling generated 43,891 unigenes with an average sequence length of 668 base pairs. A total of 26,308 (59.9%) unique sequences were annotated and 11,476 of the unique sequences were assigned to specific metabolic pathways by the Kyoto Encyclopedia of Genes and Genomes. cDNA sequences for all of the known enzymes involved in mogrosides backbone synthesis were identified from our library. Additionally, a total of eighty-five cytochrome P450 (CYP450) and ninety UDP-glucosyltransferase (UDPG) unigenes were identified, some of which appear to encode enzymes responsible for the conversion of the mogroside backbone into the various mogrosides. Digital gene expression profile (DGE) analysis using Solexa sequencing was performed on three important stages of fruit development, and based on their expression pattern, seven CYP450s and five UDPGs were selected as the candidates most likely to be involved in mogrosides biosynthesis.CONCLUSION:A combination of RNA-seq and DGE analysis based on the next generation sequencing technology was shown to be a powerful method for identifying candidate genes encoding enzymes responsible for the biosynthesis of novel secondary metabolites in a non-model plant. Seven CYP450s and five UDPGs were selected as potential candidates involved in mogrosides biosynthesis. The transcriptome data from this study provides an important resource for understanding the formation of major bioactive constituents in the fruit extract from S. grosvenorii.

Many researches had been done on genic male sterility gene,which mainly include the observation,molecular markers and mapping,gene expression,control and cloning,transgenic,producing application and routine breeding of genetic characters of male sterility materials,etc.The recent research progress were summarized,the utilization prospect and development direction were forecasted.