Campus Access Only

All rights reserved. This publication is intended for use solely by faculty, students, and staff of University of the Pacific. No part of this publication may be reproduced, distributed, or transmitted in any form or by any means, now known or later developed, including but not limited to photocopying, recording, or other electronic or mechanical methods, without the prior written permission of the author or the publisher.

Date of Award


Document Type


Degree Name

Master of Science (M.S.)


Behavioral Sciences

First Advisor

Joan Lin Cereghino

Second Advisor

Geoff Lin Cereghino

First Committee Member

Amir Assadi-Rad

Second Committee Member

Srinivas Venkatram

Third Committee Member

Craig Vierra


The methylotrophic yeast, Pichia pastoris, has been used to successfully produce and express over 700 heterologous proteins. Currently, the only two dominant selectable markers available are the Zeocin and blasticidin resistance genes. This study focused on characterizing and developing the modified G418 selectable marker so it could be used for primary selection of single copy and multicopy strains. To demonstrate its use as a selectable marker the SLPI reporter gene was inserted into the Zeocin resistance vector, pPICZB, and the new G418 resistance vector, pKANa B. The resulting constructs were then transformed into P. pastoris yJC100 cells. Piggyback strains containing both resistance vectors were also constructed. Real-time PCR, spot western blot, western immunoblot, and ELISA analysis confirmed that the modified G418 resistant vector was comparable to the Zeocin resistant vector and that its capable of generating multicopy transformants. In addition, analysis of our piggyback strains showed an increase in SIPI production and in plasmid copy number when compared to their respective "mother





Rights Statement

Rights Statement

In Copyright. URI:
This Item is protected by copyright and/or related rights. You are free to use this Item in any way that is permitted by the copyright and related rights legislation that applies to your use. For other uses you need to obtain permission from the rights-holder(s).