The Biological Effects Of Yeast Strain KH8by Nitrogen Ion Implantation

Ion beam irradiation is a new and exciting method for creating transgenicorganisms, and ion beam-induced exogenous DNA transfer is also a new method formolecular hybridization between different species. So, the paper investigated a newred compound-producing recombined strain of6076with thin-layer chromatography(TLC) analysis, which was obtained by random transformation of Glycyrrhizauralensis genomic DNA mediated by nitrogen ion irradiation into Candidaguilliermondii KH8. With the aim of investigating the new red compound-producingmechanisms of ion irradiation, we chose the6076strain as target, and combined atwo-dimensional fluorescence difference gel electrophoresis (2-D DIGE) approachwith mass spectrometry and suppression subtractive hybridization (SSH) to compareprotein expression alterations and differentially expressed genes between therecombined strain and the original strain.In the recombined strain6076, we found57proteins differentially expressedwhen cultured in liquid medium for96h compared with the original strain. Of these,26protein spots were upregulated and30were downregulated. In total,56proteinspots were identified by MALDI-TOF and tandem (TOF-TOF) mass spectrometry. Bybioinformatics analysis, it could be seen that the main functions of differentiallyexpressed proteins were carbohydrate and energy metabolic process, proteinmetabolism, cell signaling, nucleotide biosynthesis, salvage and stress response, andsome proteins function were unknown. The protein score confidence interval ofprotein profiling in the down-regulated protein spots273,1249,1294and1347were81.371%,99.563%,12.864%and99.986%, respectively. Among these, the proteinscore CI of the protein spots273and1294were81.371%and12.864%. Thus, webelieve that the protein score CI is less than85%for this model organism, and that thesearch results were unauthentic. This was probably due to the irradiation by nitrogenions contributing to the mutation of these two proteins. Their lower expression may block the original metabolic pathway and thus lead to the accumulation of the new redcompound.On the other hand, a suppression subtractive hybridization (SSH) technology wasused to construct a subtractive cDNA library of recombined strain6076. Throughsequencing analysis, from a forward subtractive cDNA library a total of14genefragment sequences with high expression levels were obtained, which included9differentially expressed genes. Among these genes,7differentially expressed genesrevealed the mutation in bases. Meanwhile, among these7differentially expressedgenes,4genes with base mutation resulted in the change of protein sequences. Inaddition, the base deletion was also observed in one gene fragment sequence. Afterthe predicted protein sequence was compared with its homologous proteins, itsfunctions were still unclear. And from a forward subtractive cDNA library of theoriginal strain KH8,10fragment sequences with highly expressed genes wereobtained. Through analyzing the isolated differentially expressed genes involved inmetabolic pathways, and some enzymes were highly expressed in recombined strain6076after96h cultivation, the transgenic yeast after96h cultivation still revealedstronger metabolic capability than original strain.The biological effects of yeast strain KH8by the nitrogen ion implantation werestudied by the methods of proteomics and differentially expressed genes. The resultsmade staggered, and some important conclusions had obtained. However, thedifferentially expressed proteins and differentially expressed genes and therelationship between the metabolism of red materials is still needs further research.