Clone distribution / genome library screening
The Leiden Genome Technology Center (LGTC) originally started as the YAC Screening Centre Leiden (YSCL). After initial national funding through Nederlandse organisatie voor Wetenschappelijk Onderzoek (NWO), the YSCL participated in a 4-year EU-funded consortium project originated to acquire, maintain and distribute large insert DNA clone resources throughout Europe. The contribution of the LGTC to this effort was to maintain libraries, to distribute clones, to perform library screening (by hybridization and / or PCR) and to exchange knowledge and protocols. The LGTC facilities have been continuously expanded and included YAC, PAC and specialized cosmid libraries, including chromosome specific and Fugu cosmid libraries.
The funding to offer these services for free ended in July 1996. The LGTC decided to continue its activities and finance them through a ‘fee-for-service’ system. In this set-up the LGTC is not commercial but rather aims to keep its services active on a non-profit basis.
DNA-chip and micro-array technology
The Affymetrix DNA-chip and micro-array technology at the LGTC was established as a joint effort of the Center for Human and Clinical Genetics (Prof. Dr. Gert-Jan. van Ommen / Dr. Johan den Dunnen) and Molecular Cell Biology (Prof. Dr. Hans Tanke / Prof. Dr. Ton Raap), with financial support from NWO and the LUMC. Additional support was obtained from the Centre for Biomedical Genetics (CBG) to establish the DNA-chip array technology in the setting of the CBG. From 2003 onward, the array technology is supported by the Center for Medical Systems Biology (CMSB), a Netherlands Genomics Center of Excellence established by the Netherlands Genomics Initiative (NGI). Within the CMSB the LGTC acts for the LUMC as the Systems Biology array technology platform. Through this support the LGTC has been able to upgrade the systems presents to the highest possible standards.
In January 2009 the LGTC purchased the FlexArrayer from FlexGen, a spin-off from Dutch Space and the Leiden University Medical Centre. The joint efforts of FlexGen and the LUMC made the development of the FlexArrayer (a system to synthesize custom arrays) possible.
SNP typing & Mutation Detection
In 2005 the LGTC acquired an Illumina BeadArray system and in 2007 the Illumina BeadXpress system, facilitating high-throughput SNP-typing, both targeted and genome-wide. In addition, the flexibility of the system facilitates its use for genome-wide gene expression profiling. High-throughput SNP-typing is further supported by an ABI 7900HT system (TaqMan), acquired in 2005 and a Roche Lightcycler 480 acquired in 2006. SNP-discovery and mutation detection is facilitated both by our sequencing possibilities (see above) as well as through the availability of a LightScanner (Idaho Technology) for high-resolution Melting Curve Analysis (HRMA). In 2008, as the first European lab, the LGTC acquired the Fluidigm Biomark real-time PCR machine. In 2012 the Roche LightCycler 480 was replaced by a LightCycler 480 V2.
In 1992 the LGTC initiated an internal Sequencing Core facility by employing an Automated Laser Fluorescence DNA-sequencer (ALF), one of the first automated DNA sequencing devices in Nederland. With increasing demand and the improvement of big-dye terminator chemistry, an ABI-377 slab gel system replaced the previous system in 1996. At the same time the sequencing services were extended beyond the borders of the LGTC. In 1999 the ABI PRISM 3700 DNA analyzer, a fully automated 96-capillary electrophoresis instrument, designed for production-scale DNA analysis, was purchased to meet the increasing demand for both sequence and fragment analysis. In 2002 this system was extended with an ABI 3730 DNA analyzer and replaced by an ABI 3730xl 96-cappilary system in 2008. Today, the LGTC is the Sequencing Core facility for the LUMC.
To diversify its sequencing possibilities, the LGTC in 2000 implemented “Pyrosequencing” technology. Later, after the introduction of DNA-chip and micro-array technology, array-based re-sequencing was introduced. In 2007, supported by the Centre for Biomedical Genetics (CBG), the LGTC acquired a mega-base sequencing systems, the Genome Analysis System from Solexa/Illumina. In 2008 the LGTC aquired a second Illumina Genome Analysis System and soon after both systems have been completed with Paired-End modules and upgraded to GAIIx analyzers.
Next Generation Sequencing
It soon became clear that the focus of the LGTC should be in next generation sequencing and in December 2009 the LGTC acquired Helicos single cell sequencer. It was followed by an Illumina HiSeq2000 in January 2010. In general, technology is available from two or more suppliers giving the possibility to make the best choice based on the combination of performance and cost. To extend the possibilities of next generation sequencing an Ion Torrent PGM from Life Technologies was purchased in April 2011 and a Pacific Biosciences RS, a real time single molecule sequencer was acquired in December 2011. With the increase of projects and sample preparation the LGTC implemented a Caliper NGSx robot in 2012, for automated library preparations as well as isolation and purification of DNA and RNA and PCR fragments.
Unfortunately Helicos bankrupted in December 2012 but we are still looking at options to repair our system. We have collaborations with the TU Delft and with the technical department of the LUMC to explore the possibilities.
In October 2013 the LGTC acquired an Ion Proton from Life Technologies and at the same time a Fluidigm C1 Single-Cell Auto prep system was purchased. The focus is switching much more to analyzing the DNA and RNA of a single cell and the Fluidigm C1 should broaden the possibilities.
To expand it’s capacity and the turn around time of projects a HiSeq2500 and a NextSeq500 from illumina have been purchased in 2014.