isolation-viable-multicellular-glands-from-tissue-carnivorous-plant

Isolation of Viable Multicellular Glands from Tissue of the Carnivorous Plant, <i>Nepenthes</i>

Many plants possess specialized structures that are involved in the production and secretion of specific low molecular weight compounds and proteins. ...

Université de Lorraine

A rapid mechanical micropreparation technique has been developed to isolate multicellular glands, here from Nepenthes pitchers, based on a microdissection platform. The method is an alternative to laser capture dissection because fresh plant tissue can be used directly without previous fixation. Subsequent experiments, such as polymerase chain reaction (PCR)-based detection of an individual gene encoding a thaumatin-like protein and RNA extraction for gene expression analysis, have been successfully added to prove the quality of the prepared biological material. The procedure described is adaptable to a broad range of plant species and should find wide application in the preparation of multicellular glands or other tissues.

Micropreparation of single secretory glands from the carnivorous plant Nepenthes.

Carnivory in plants is an adaptation strategy to nutrient-poor environments and soils. Carnivorous plants obtain some additional mineral nutrients by trapping and digesting prey; the genus Nepenthes is helped by its specialized pitcher traps. To make the nutrients available, the caught prey needs to be digested, a process that requires the concerted activity of several hydrolytic enzymes. To identify and investigate the various enzymes involved in this process, fluid from Nepenthes traps has been analysed in detail. In this study, a novel type of Nepenthes endochitinase was identified in the digestion fluid of closed pitchers. The encoding endochitinase genes have been cloned from eight different Nepenthes species. Among these, the deduced amino acid sequence similarity was at least 94.9%. The corresponding cDNA from N. rafflesiana was heterologously expressed, and the purified protein, NrChit1, was biochemically characterized. The enzyme, classified as a class III acid endochitinase belonging to family 18 of the glycoside hydrolases, is secreted into the pitcher fluid very probably due to the presence of an N-terminal signal peptide. Transcriptome analyses using real-time PCR indicated that the presence of prey in the pitcher up-regulates the endochitinase gene not only in the glands, which are responsible for enzyme secretion, but at an even higher level, in the glands' surrounding tissue. These results suggest that in the pitchers' tissues, the endochitinase as well as other proteins from the pitcher fluid might fulfil a different, primary function as pathogenesis-related proteins.

Functional characterization of a class III acid endochitinase from the traps of the carnivorous pitcher plant genus, Nepenthes.

Carnivorous plants of the genus Nepenthes possess modified leaves that form pitfall traps in order to capture prey, mainly arthropods, to make additional nutrients available for the plant. These pitchers contain a digestive fluid due to the presence of hydrolytic enzymes. In this study, the composition of the digestive fluid was further analysed with regard to mineral nutrients and low molecular-weight compounds. A potential contribution of microbes to the composition of pitcher fluid was investigated.

Article	Year
Micropreparation of single secretory glands from the carnivorous plant Nepenthes. Analytical biochemistry\| PubMed ID: 19602419	2009
Functional characterization of a class III acid endochitinase from the traps of the carnivorous pitcher plant genus, Nepenthes. Journal of experimental botany\| PubMed ID: 21633084	2011
Secreted pitfall-trap fluid of carnivorous Nepenthes plants is unsuitable for microbial growth. Annals of botany\| PubMed ID: 23264234	2013

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