@masterthesis{Giebel, type = {Bachelor Thesis}, author = {Giebel, Alice}, title = {Cloning Of Human Secretory Gelsolin In Pichia Pastoris}, url = {http://nbn-resolving.de/urn:nbn:de:hbz:1383-opus4-6585}, school = {Hochschule Rhein-Waal}, pages = {74}, abstract = {The production of recombinant proteins plays an important part in biotechnology for over 30 years. Their applications range from pharmaceutical practice up to industrial (non-medical) purposes and, therefore, show great importance in today´s life. Most recombinant proteins are being produced by genetic modification of microorganisms to either generate those intracellularly or to secrete the desired protein into their surroundings. Dependent of the expression mechanism, the production can be influenced and improved to achieve high yields and protein purity. One aim of this study was to determine and optimise the most favorable cloning strategy for the expression and secretion of the plasma form of the actin binding protein gelsolin from Pichia pastoris. Gelsolin has two isoforms of which one of them gained particular attention due to its application in research projects concerning the current COVID-19 pandemic. For this project, a variety of oligonucleotide primers were designed to amplify the whole sequences and various parts of both gelsolin isoforms during PCR approaches. The accuracy and efficacy of primers is determined by several factors such as their length and exact structure. Also, many factors depending on the sequence of interest are influencing the outcome of the PCR. Therefore, the second aim of this study was to design and validate primer pairs with most preferable conditions to amplify the sequence for the expression of gelsolin and optimise the PCR conditions for this purpose.}, language = {en} }