Expression of Acyl-lipid 12-desaturase Gene in Prokaryotic and Eukaryotic Cells and Its Effect on Cold Stress Tolerance of Potato

We report the expression profile of acyl-lipid △12-desaturase （desA） gene from Synechocystis sp. PCC6803 and its effect on cell membrane lipid composition and cold tolerance in prokaryotic （Escherichia coil） and eukaryotic （Solanum tuberosum） cells. For this purpose, a hybrid of desA and reporter gene encoding thermostable lichenase （licBM3） was constructed and used to transform these cells. The expression of this hybrid gene was measured using qualitative （Petri dish test, electrophoregram and zymogram） and quantitative methods （spectrometry and gas liquid chromatography assays）. The maximum level of linoleic acid in the bacterial cells containing hybrid gene was 1.9% of total fatty acids. Cold stress tolerance assays using plant damage index and growth parameters showed that cold tolerance was enhanced in primary transgenic lines because of increased unsaturated fatty acid concentration in their lipids. The greatest content of 18：2 and 18：3 fatty acids in primary transgenic plants was observed for lines 2 （73%） and 3 （41%）. Finally, our results showed that desaturase could enhance tolerance to cold stress in potato, and desaturase and lichenase retain their functionality in the structure of the hybrid protein where the enzymatic activity of target gene product was higher than in the case of reporter lichenase gene absence in the construction.

Employing pH-responsive RNA triplex to control CRISPR/Cas9-mediated gene manipulation in mammalian cells

The clustered regularly interspersed short palindromic repeats/CRISPR-associated protein 9(CRISPR/Cas9)system is an RNA-guided platform for highly efficient and specific genome targeting in diverse organisms,which has been exploited for various applications in gene manipulation.Compared with the constantly active CRISPR/Cas9 function,conditional control of its activity can improve the performance of the system with reduced side effects and high spatiotemporal precision.The pH-responsive triplex RNA was successful used in CRISPR-derived RNA/trans-activating crRNA(crRNA/tracrRNA)of CRISPR/Cas9,thus affecting RNA/dead Cas9(dCas9)complex to target DNA in vitro and in vivo.This design of triplex RNA opens a new window towards the broad involvement of eukaryotic cells for conditional control of CRISPR/Cas9function.?2024 Published by Elsevier B.V.on behalf of Chinese Chemical Society and Institute of Materia Medica,Chinese Academy of Medical Sciences.

IRESfinder:Identifying RNA internal ribosome entry site in eukaryotic cell using framed k-mer features

In eukaryotic cells, initiation of protein translation is to recruit the ribosome to a specific mRNA, which is generally dependent on the 5＇ cap structure. However, protein translation can also be initiated in a cap-independent manner by using a cis-regulatory element termed the internal ribosome entry site （IRES）. The first experimentally validated IRES was reported in the poliovirus （Pelletier and Sonenberg, 1988）. Then eukaryotic cellular mRNAs were also validated to contain IRES elements.