Cloning and Expression Analysis of Polyubiquitin Gene (LcUBQ) in Luffa cylindrica

Abstract: Ubiquitin is a stress responses protein which mediates ubiquitin/26S proteasome pathway and plays an important role in plant adaptation to various environmental stresses. To investigate the function of polyubiquitin gene in Luffa cylindrica， LcUBQ was isolated from luffa by using RACE (rapid-amplification of cDNA ends) and RT-PCR (reverse transcription-PCR) techniques (GenBank accession number was JN979372). It was 1 579 bp， which contained a 1 371 bp open reading frame (ORF) that encoded 457 amino acids， with a predicted molecular weight of 51.26 kDa and a hypothetical isoelectric point of 7.05. This protein lacked the signal peptides and membrane-spanning domains and the WoLF PSORT protection indicated that it was located in cytoplasm. The secondary and tertiary protein structures were predicted， which consisted of 24.73% α-helices， 22.54% extended strands， and 52.74% random coils. Phylogenetic analysis illustrated that LcUBQ had high similarity to the UBQ of Cucurbitaceae plants such as Cucumis sativus and Cucumis melo. The results of fluorescence quantitative PCR analysis revealed that LcUBQ was expressed in roots， stems， leaves， flowers and fruits， and the lowest expression was found in roots and leaves. LcUBQ could be induced in leaves by high temperature (40 °C)， low temperature (4 °C) and low light (20 μmol/(m2∙s)) stresses， especially low temperature stress. Under low temperature stress， the expression of LcUBQ presented as an increased at first and then decreased trend， and the trend was consistent with the changes in the corresponding ubiquitin protein levels. Moreover， the levels of LcUBQ and ubiquitin protein were markedly elevated after stress for 2 h whether low temperature or low light. The results suggested that LcUBQ might be involved in early stress signal transduction and low temperature and low light stress response process in Luffa cylindrica.

CSTR: 32200.14.cjcb.2018.01.0012