The Max-Planck-Institute of Molecular Genetics Mouse Genome Project


Functional characterisation of the mouse genome requires the availability of a comprehensive physical map to obtain molecular access to chromosomal regions of interest. Positional cloning remains a crucial way of linking phenotype with particular genes. A key step and frequent stumbling block in positional cloning is making a contig of a genetically defined candidate region. The most efficient first step is isolating YAC (Yeast Artificial Chromosome) clones. A robust, detailed YAC contig map is thus an im portant tool. We here present the physical map of the mouse genome that combines the results of our group at The Max-Planck-Institute of Molecular Genetics (MPI-MG) with the mouse map data generated at the Whitehead Institute for Biomedical Research/MIT Center for Genome Research (WICGR), recently published by Nusbaum et al and accessible online. Employing Interspersed Repetitive Sequence (IRS)-PCR genomics, we have generated an advanced second-generation YAC contig map of the mouse genome which doubles both the depth of clones and the density of markers available. In addition to the primarily YAC -based map, we located 2224 BAC (Bacterial Artificial Chromosome) clones. This allows us to present for the first time a dense framework of BACs spanning the genome of the mouse which for instance can serve as a nucleus for genomic sequencing. Four large- insert mouse YAC libraries from three different strains are included in our data and our analysis incorporates the data of Hunter et al. (1996) and Nusbaum et al. (1999). There is a total of 21904 markers on the final map, 12136 from our own data, and a t otal of 50891 YACs, of which 32773 are positive for more than one marker.

Go to chromosome maps

The interspersed repetitive sequence (IRS)-PCR technology is based on the abundance of repeat elements in the genome of higher organisms. Repetitive sequence primers are used to amplify sequences that are flanked by repeat elements. For instance, a single primer to a portion of the B1 repeat will amplify thousands of individual fragments from a mouse genomic DNA template. IRS-PCR can be used on any genomic DNA containing sample, i.e. total genomic DNA, DNA from cell hybrids, from individual clones or clon e pools. Complex IRS-PCR reaction products can be cloned into plasmids. An alternative, which we used extensively, is IRS-PCR on YAC- or BAC clones. Fragments generated on these low-complexity templates can directly be employed as markers. The generation of large numbers of IRS-markers in this way is cheap, since there is no requirement to sequence markers or to design locus-specific primers. For this work, IRS-PCR probe fragments were generated with a single B1 repeat-derived primer from the following so urces:
 
 

Library/probe identification Total number of probes on map Probe origin cell line---strain origin
mbacr 2215 CITB Mouse BAC library (B. Birren, unpublished) 129/Sv (ES cell line CJ7)
bir 1247 IRS-PCR fragment library generated from total genomic DNA C57BL/6
173r 176 IRS-PCR fragment library generated from cell hybrid line 167 EJ 167EJ
whtII 8395 Mouse YAC clones from Haldi et al. (1996) library C57BL/6

Table 1: MPIMG-markers

Mapping reagents such as genetic crosses, radiation hybrid panels and genomic libraries can be typed with IRS-PCR markers by hybridisation. The target DNA (complex IRS-PCR products generated from the mapping reagent) is arrayed at high density onto a mem brane and probed with one labelled marker fragment at a time. Each of the IRS-probes that we generated was hybridised against filters containing IRS-PCR products generated from YAC pools, representing the four available large insert mouse YAC libraries (7 5 000 YAC clones in total) constructed at the ICRF and the Whitehead Institute:
 
 
Library/clone identification Number of clones Genome coverage strain of origin Reference
ICRFy902 13 400 2 x C3H Larin et al., 1993
ICRFy903 5 000 0.8 x C57BL/10 Larin et al., 1993
WHTy910 20 000 4.5 x C57BL/6 Kusumi et al., 1993
whtII (WHTy917) 40 800 13 x C57BL/6 Haldi et al., 1996

Table 2: Mouse YAC libraries


Our advanced mouse physical map integrates our own data with the results of two mouse mapping projects from Nusbaum et al. (1999) and Hunter et al. (1996). This maximizes the impact of the map produced, as it allows for easy map comparison to be conducted. Specifically, the following markers and data were integrated:
 
 
Source Marker type Marker designation Number of probes on map Reference
WICGR SSLP-STS D*Mit* 4314 Nusbaum et al., 1999
WICGR STS Various 3534 Nusbaum et al., 1999
K. Hunter IRS-PCR marker HUN 324 Hunter et al., 1996

Table 3: Outside data integrated into the map

 
 

All markers generated by us are publicly available from the Resource Center of the German Human Genome Project (RZPD). Likewise, all large-insert clone libraries (BACs, YACs) that we used for data production and map generation are from public sources. Ind ividual clones can be ordered from various genome centers, including RZPD in Berlin. In this context it should be noted that for any BAC- and YAC-derived probe, markers can be regenerated with IRS-PCR on the respective clone with primer B1R.
 
 

YAC clones Distributors
ICRFy903 RZPD
ICRFy902 RZPD
WHTy910 RZPD, HGMP , Research Genetics
whtII (WHTy917) RZPD, HGMP , Research Genetics
MPIMG-markers Distributors Comment
mbacr Research Genetics Plate coordinates of mbacr probes are identical with clone coordinates in CITB Mouse BAC library. IRS-PCR on colony material or BAC-DNA regenerates mbacr marker.
bir RZPD IRS-PCR fragment cloned into plasmid vector
173r RZPD IRS-PCR fragment cloned into plasmid vector
whtII (WHTy917) RZPD, HGMP , Research Genetics Plate coordinates of whtII probes are identical with clone coordinates in whtII (WHTy917) YAC library. IRS-PCR on colony material or YAC-DNA regenerates whtII marker

Table 4: Genome Centers and Institutions that distribute MPI-MG markers and YAC clones that we have placed on our maps