Mogrosides and steroid saponins are tetracyclic triterpenoids found in Siraitia grosvenorii.Squalene synthase(SQS) and cycloartenol synthase(CAS) are key enzymes in triterpenoid and steroid biosynthesis.In this study,...Mogrosides and steroid saponins are tetracyclic triterpenoids found in Siraitia grosvenorii.Squalene synthase(SQS) and cycloartenol synthase(CAS) are key enzymes in triterpenoid and steroid biosynthesis.In this study,full-length cDNAs of SgSQS and SgCAS were cloned by a rapid amplification of cDNA-ends with polymerase chain reaction(RACE-PCR) approach.The SgSQS cDNA has a 1254 bp open reading frame(ORF) encoding 417 amino acids,and the SgCAS cDNA contains a 2298 bp ORF encoding 765 amino acids.Bioinformatic analysis showed that the deduced SgSQS protein has two transmembrane regions in the C-terminal.Both SgSQS and SgCAS have significantly higher levels in fruits than in other tissues,suggesting that steroids and mogrosides are competitors for the same precursors in fruits.Combined in silico prediction and subcellular localization,experiments in tobacco indicated that SgSQS was probably in the cytoplasm or on the cytoskeleton,and SgCAS was likely located in the nucleus or cytosol.These results will provide a foundation for further study of SgSQS and SgCAS gene functions in S.grosvenorii,and may facilitate improvements in mogroside content in fruit by regulating gene expression.展开更多
Messenger RNA-like non-coding RNAs （mlncRNAs） are a newly identified group of non-coding RNAs （ncRNAs） that may be involved in a number of critical cellular events. In this study, 93 candidate porcine mlncRNAs wer...Messenger RNA-like non-coding RNAs （mlncRNAs） are a newly identified group of non-coding RNAs （ncRNAs） that may be involved in a number of critical cellular events. In this study, 93 candidate porcine mlncRNAs were obtained by computational prediction and screening, among which 72 were mapped to the porcine genome. Further analysis of 8 representative candidates revealed that these mlncRNA candidates are not highly conserved among species. Remarkably, one of the candidates, sTF35495, was found to be precursor of a putative porcine microRNA. By RACE PCR, we determined that the full length of sTF35495 was 3 kb. The protein-coding potential of this RNA was tested in silico with no significant finding. Semi-quantitative RT-PCR analysis of the subgroup of 8 candidates revealed two distinct expression profiles and two molecules were further validated by real-time PCR. The predicted pre-microRNA sequence in this study provides a potentially interesting insight into the in vivo function of porcine mlncRNAs and our findings suggest that they play key biological roles in Sus scrofa.展开更多
Salt and drought limit the range of applications of perennial ryegrass(Lolium perenne L.), which is one of the important turf and forage grasses. Previous studies have suggested that pyrroline-5-carboxylate reductase(...Salt and drought limit the range of applications of perennial ryegrass(Lolium perenne L.), which is one of the important turf and forage grasses. Previous studies have suggested that pyrroline-5-carboxylate reductase(P5CR) might play a central role in proline accumulation in plants that are responsive to stresses. In the present study, the Lolium perenne L. pyrroline-5-carboxylate reductase(LpP 5CR) gene was cloned from leaves of the cultivar ‘Derby' using the RACE technique. The full-length LpP 5CR gene was 1 047 bp in length, which comprised an open reading frame(ORF) of 840 bp in size. Sequence alignment revealed that the putative Lp P5 CR had a 94.3% similarity to Ta P5 CR. q RT-PCR displayed that the mR NA levels of the LpP 5CR gene were almost the same as that in the roots, stems, and leaves of perennial ryegrass seedlings subjected to normal condition or NaC l treatment for 1 h. Moreover, the transcription level of LpP 5CR was up- or down- regulated with Na Cl, polyethylene glycol(PEG), cold, or abscisic acid(ABA) treatment for 3 to 48 h. In addition, confocal microscopy localized the GFP-Lp P5 CR fusion protein to the cytoplasm of onion epidermal cells. These findings suggest that LpP 5CR encodes a cytoplasmic P5 CR protein that plays an important role in the response of perennial ryegrass to various stresses.展开更多
基金supported by the National Nature Science Foundation of China(No.81373914)National Key Technology Support Program(No.2011BAI01B03)Guangxi Natural Science Fundation(Nos.2013GXNSFDA019021 and 2012GXNSFAA053043)
文摘Mogrosides and steroid saponins are tetracyclic triterpenoids found in Siraitia grosvenorii.Squalene synthase(SQS) and cycloartenol synthase(CAS) are key enzymes in triterpenoid and steroid biosynthesis.In this study,full-length cDNAs of SgSQS and SgCAS were cloned by a rapid amplification of cDNA-ends with polymerase chain reaction(RACE-PCR) approach.The SgSQS cDNA has a 1254 bp open reading frame(ORF) encoding 417 amino acids,and the SgCAS cDNA contains a 2298 bp ORF encoding 765 amino acids.Bioinformatic analysis showed that the deduced SgSQS protein has two transmembrane regions in the C-terminal.Both SgSQS and SgCAS have significantly higher levels in fruits than in other tissues,suggesting that steroids and mogrosides are competitors for the same precursors in fruits.Combined in silico prediction and subcellular localization,experiments in tobacco indicated that SgSQS was probably in the cytoplasm or on the cytoskeleton,and SgCAS was likely located in the nucleus or cytosol.These results will provide a foundation for further study of SgSQS and SgCAS gene functions in S.grosvenorii,and may facilitate improvements in mogroside content in fruit by regulating gene expression.
基金supported by the National Natural Science Foundation of China (No.30830080 and 30800779)the Natural Science Foundation of Beijing (No.5072035)the Ministry of Science and Technology of China (No.2006AA10Z135,2008AA10Z143,2006CB102105 and 2009CB941604)
文摘Messenger RNA-like non-coding RNAs （mlncRNAs） are a newly identified group of non-coding RNAs （ncRNAs） that may be involved in a number of critical cellular events. In this study, 93 candidate porcine mlncRNAs were obtained by computational prediction and screening, among which 72 were mapped to the porcine genome. Further analysis of 8 representative candidates revealed that these mlncRNA candidates are not highly conserved among species. Remarkably, one of the candidates, sTF35495, was found to be precursor of a putative porcine microRNA. By RACE PCR, we determined that the full length of sTF35495 was 3 kb. The protein-coding potential of this RNA was tested in silico with no significant finding. Semi-quantitative RT-PCR analysis of the subgroup of 8 candidates revealed two distinct expression profiles and two molecules were further validated by real-time PCR. The predicted pre-microRNA sequence in this study provides a potentially interesting insight into the in vivo function of porcine mlncRNAs and our findings suggest that they play key biological roles in Sus scrofa.
基金The National High Technology Research and Development Program of China(Grant No.2011AA100209)Jilin Provincial Department of Education(Grant No.2013-23)supported this study
文摘Salt and drought limit the range of applications of perennial ryegrass(Lolium perenne L.), which is one of the important turf and forage grasses. Previous studies have suggested that pyrroline-5-carboxylate reductase(P5CR) might play a central role in proline accumulation in plants that are responsive to stresses. In the present study, the Lolium perenne L. pyrroline-5-carboxylate reductase(LpP 5CR) gene was cloned from leaves of the cultivar ‘Derby' using the RACE technique. The full-length LpP 5CR gene was 1 047 bp in length, which comprised an open reading frame(ORF) of 840 bp in size. Sequence alignment revealed that the putative Lp P5 CR had a 94.3% similarity to Ta P5 CR. q RT-PCR displayed that the mR NA levels of the LpP 5CR gene were almost the same as that in the roots, stems, and leaves of perennial ryegrass seedlings subjected to normal condition or NaC l treatment for 1 h. Moreover, the transcription level of LpP 5CR was up- or down- regulated with Na Cl, polyethylene glycol(PEG), cold, or abscisic acid(ABA) treatment for 3 to 48 h. In addition, confocal microscopy localized the GFP-Lp P5 CR fusion protein to the cytoplasm of onion epidermal cells. These findings suggest that LpP 5CR encodes a cytoplasmic P5 CR protein that plays an important role in the response of perennial ryegrass to various stresses.
