I denne masteroppgaven har det blitt utviklet primerpar for 17 humane PLA2-isotyper. Ved hjelp av primerene har vi påvist uttrykk av 17 PLA2-isotyper i HaCaT-keratinocytter, hvorav 11 av disse representerer førstegangspåvisninger i HaCaT. Funnene representerer økt kunnskap om PLA2-familien og viser at HaCaT uttrykker et mangfold av PLA2-isotyper. Primerene ble også benyttet til å kartlegge PLA2-uttrykk gjennom keratinocytters differensiering. Resultatene viser at 13 av 17 isotyper uttrykkes gjennom hele differensieringsprosessen, som indikerer at PLA2-enzymer spiller sentrale roller i lipidmetabolisme i human epidermis. PLA2G2A viste, som den eneste av de studerte isotypene, sterk oppregulering under differensieringen, som vitner om at enzymet kan spille en viktig rolle i de øvre strata i epidermis.Sammendrag Under utviklingen av primerparene ble human postnatal placenta tatt i bruk som antatt positiv kontroll for uttrykk av samtlige PLA2-isotyper. 8 av 17 primerpar detekterte mRNA i placenta, hvorav PLA2 gruppe 2D, 4C, 4D og 10 representerer førstegangspåvisninger i human placenta. Sammendrag Primerene utgjør et verktøy som gjør det mulig å kartlegge PLA2-uttrykk i alle humane celler og vev. Da diverse PLA2-isotyper har blitt vist å spille en sentral rolle i inflammasjonssykdommer ved å katalysere hydrolysen av arakidonsyre fra glyserofosfolipider, vil det være intressant å kartlegge PLA2-uttrykk i friskt vev versus betent vev. Primerene vil legge grunnlaget for økt forståelse av de mange PLA2-enzymenes funksjoner i inflammasjonssammenheng, og potensielt legge grunnlag for utvikling av målrettede medisiner mot kronisk inflammatoriske sykdommer.

IDL6 and IDL7 are postulated peptides of the INFLORESCENCE DEFICIENT IN ABSCISSION (IDA) peptide family in the model organism Arabidopsis thaliana. The genes encoding the postulated peptides, IDL6 and IDL7, were investigated to potentially elucidate the function of the peptides and the possible relations to plant defense and stress tolerance in A. thaliana. A phenotypic characterization study of single and double knockout mutant lines, and over-expression lines of IDL6 and IDL7 was conducted to potentially find a phenotype linked to the genes by analyzing deviations in growth and development, compared to wild type (Wt) A. thaliana. Over-expression lines showed a tendency to have a higher amount of individuals reaching defined growth stages in seedling development, but this could not be concluded to be a phenotype linked to IDL6 or IDL7.The genes have been shown to be highly up-regulated in response to several different stress treatments, both abiotic and biotic in silico data. Several different transgenic lines of A. thaliana were subjected to different types of stress treatments to potentially verify the in silico data. Knockout lines, double knockout lines, over-expression lines and promoter:GUS lines of IDL6 and IDL7 were treated with abiotic stress factors (NaCl, mannitol, UV-B light, H2O2 and paraquat) and biotic stress factors (aphids, Pseudomonas syringae and flagellin22) to investigate differences in tolerance, gene expression and promoter activity in the respective transgenic lines and Wt A. thaliana.No absolute phenotype was detected in experiments related to stress tolerance, and great variations in promoter activity using promoter:GUS lines were observed. However, double knockout mutants of idl6 and idl7 showed a small trend to tolerate NaCl better than Wt A. thaliana and other transgenic lines. The great variation in GUS expression from GUS assay lead to a thorough screening and expression analyses of promoter:GUS lines. Results presented in this work, indicate that expression of the IDL6 and IDL7 genes may be subjected to extensive post transcriptional regulation through mRNA degradation, possibly governed by stress related environmental signals. A novel member of the IDA peptide family, IDL8, was also analyzed. Segregation analyses of knock out lines were conducted and they were genotyped to verify that mutant lines of idl8 were real knockouts. Results presented here show that one of the idl8 mutant lines had the T-DNA inserted to the promoter region of the gene and is postulated to be a real knockout. However, further expression analyses should be conducted to verify that the gene is not transcribed. Over-expression lines of IDL8 were successfully constructed through recombinant DNA technology and by T-DNA insertion using Agrobacterium tumefaciens.

This thesis describes the cytochrome p450 superfamily in the pennate diatom Seminavis robusta. This diatom has a benthic way of life and can adhere to surfaces, and is a new candidate model organism representing the pennate diatoms. The genome of S. robusta is currently being sequenced and is characterized. This revealed that the genome of S. robusta contains 68 genes encoding CYPs, an unusually high number compared to other related organisms. Several unique families and subfamilies were discovered, but also known families from both plants and animals. This reflects the diversity of factors that may have contributed to the evolution of these genes in diatoms, and possibly how a benthic lifestyle may drive this process.Both bioinformatical tools and transcriptional analysis have been used to characterize and explore this superfamily. An experimental study was performed by cultivating S. robusta at different conditions, including normal temperature and light, lower temperature and a day/night cycle. This confirmed some of the CYPs’ involvement in light-/dark-responses, as well as some responses that seemed coupled to temperature.As part of the study, a comparison was made between two software programs used for analyzing gene expression data. This concluded that replacing the software currently used with a new software program with more possibilities will be beneficial for ensuring a higher quality assurance when publishing RT-qPCR data, as well as one step closer publishing according to the MIQE-guidelines.