Mouse Genetics: Concepts & Applications (Full Table of Contents)

Copyright ©1995 Lee M. Silver

Suppliers of Mice

A.1 Major suppliers of common inbred and outbred strainsa

A.2 Other commercial sources of micea

A.1 Major suppliers of common inbred and outbred strainsa

SUPPLIER
SERVICES
ADDRESS
CONTACT NUMBERS
Charles River Laboratories
Established strains, transgenics
251 Ballardvale St.

Wilmington, MA 01887

Phone: 508-658-6000

FAX: 508-658-7132

Harlan Sprague Dawley, Inc.
Established strains, transgenics
P.O. Box 29176

Indianapolis, IN 46229

Phone: 317-894-7521

FAX: 317-894-4473

Jackson Laboratory
Established strains
Animal Resources

600 Main St.

Bar Harbor, ME 04609-0800

Phone: 800-422-6423

FAX: 207-288-3398

Taconic Farms, Inc.
Established strains, transgenics
33 Hover Avenue

Germantown, NY 12526

Phone: 518-537-6208

FAX: 518-537-7287

aListed in alphabetical order

A.2 Other commercial sources of micea

SUPPLIER
SERVICES
ADDRESS
CONTACT NUMBERS
Baekon, Inc.
Transgenics
4420 Enterprise St.

Fremont, CA 94538

Phone: 510-683-8881

FAX: 510-683-8712

GenPharm Intl.
Transgenics
2375 Garcia Ave.

Mountain View, CA 94043

Phone: 415-964-7024

FAX: 415-964-3537

Hilltop Lab Animals, Inc.
Established strains
Hilltop Drive

Scottsdale, PA 15683

Phone: 800-245-6921

FAX: 412-887-3582

Life Sciences, Inc.
Established strains, transgenics
2900 72nd St., North

St. Petersburg, FL 33710

Phone: 800-237-4323

FAX: 813-347-2957

aListed in alphabetical order

Appendix B

Computational Tools and Electronic Databases

B.1 On-line electronic databases

B.1.1 Mouse genetics

B.1.2 Humans and other organisms

B.2 Mouse Genetics Community Bulletin Board

B.3 Computer Programs

B.3.1 Linkage analysis

B.3.2 Mouse colony management and genetic databases

 

B.1 On-line electronic databases

An ever-increasing amount of genetic information can be easily accessed and selectively retrieved over the Internet by using a software interface protocol known as Gopher, developed at the University of Minnesota. Any researcher with an Internet link can take advantage of this protocol by loading onto their computer a platform-specific version of a "Gopher client program" available without charge from the Gopher development group. Their E-mail address is [Gopher@boombox.micro.umn.edu]. Versions of the Gopher client program are available for all common operating systems that run on IBM-compatible PCs, Macintoshs, workstations, minicomputers, and mainframes. The Gopher program takes advantage of the custom features of each local machine type and allows a user to move seemingly seamlessly among different computer locations (called Gopher Holes) around the world where database searches can be conducted and selected data can be retrieved in a simple manner.

With the addition of the NCSA Telnet protocol, direct Internet links can also be made to login-based servers on the Gopher network. Another Gopher-like tool called Fetch (developed at Dartmouth College and also available free to non-profit organizations and individuals) provides Macintosh users with the ability to download files from FTP (File Transfer Protocol) sites in a transparent manner. For more information on this program, send an E-mail message to Fetch@Dartmouth.edu.

As this book is being written, the Gopher software and its associated formats are still less than three years old and expanding rapidly. It is a near-certainty that the data sources described below will be enriched and supplemented with new sources of information and modes for their retrieval. In addition, other more sophisticated electronic highways are under development. The most advanced of these, known as the "World Wide Web (WWW)", can be accessed with the use of NCSA Mosaic client software. You should contact your local computer advisor for up-to-the-date information on the most advanced system available at the time of this reading.

B.1.1 Mouse genetics

The central database server for mouse geneticists is located at the Jackson Laboratory. It can be reached in a number of different ways: (1) By using Gopher to trace a path from the "Home Gopher Hole" at the University of Minnesota through the U.S. to Maine to "The Jackson Laboratory." (2) By pointing Gopher directly at GOPHER.INFORMATICS.JAX.ORG, (3) By using NCSA Mosaic to connect to the Jackson Laboratory through the World Wide Web. This Gopher Hole contains a number of useful databases including the Mouse Locus Catalog ( or MLC), and the Genomic Database of the Mouse (or GBASE).

MLC originated as an electronic version of the chapter entitled "Catalog of mutant genes and polymorphic loci" (Green, 1989) in the Lyon and Searle compendium Genetic Variants and Strains of the Laboratory Mouse. It is updated regularly by staff members of the Jackson Laboratory led by Dr. M.T. Davisson and Dr. D. P. Doolittle. Each locus is represented by a separate record within the database that contains a detailed description of phenotype and other characteristics. The entire database can be searched to retrieve a list of loci (with associated descriptions and citations) that contain a particular search word or combination of words anywhere within the locus record.

GBASE is a comprehensive database at the Jackson Laboratory containing mouse mapping information. Investigators must login to GBASE (as a guest) to access the data contained within it. GBASE can be reached either through Gopher or via a direct internet linkage with the NCSA Telnet protocol. Once on-line, an investigator can retrieve the accumulated linkage data published on any set of loci, linkage maps, as well as allele distribution patterns for common inbred strains and all associated citations. For further assistance, send an E-mail message to mgi-help@Jax.Org.

The Jackson Laboratory database server also has catalog files that can be downloaded with information on available mouse strains and mutant stocks (both live and in the form of DNA samples). Many other sources of genetic information, both at the Jackson Laboratory and elsewhere are also accessible through this Gopher Hole. One can jump from this Gopher Hole directly to the "Computational Biology" gopher hole with human genetic data at Johns Hopkins University (described below), and one can also search through the Genbank sequence database directly from here. It is also possible to download updated data files for use by the Encyclopedia of the Mouse Genome which is described below.

The Genome Center at the Whitehead Institute, MIT provides a highly specialized but very useful source of information through either an automated E-mail query and answer protocol or anonymous FTP (File Transfer Protocol). This Genome Center has characterized and mapped over 3,000 polymorphic mouse microsatellite markers (as of Januray, 1994) that are each defined by a unique pair of oligonucleotide primers. By filling-out and transmitting a special E-mail query form, an investigator can receive an automatic response containing information about sets of microsatellite markers defined by various criteria, including chromosomal location and polymorphisms between particular strains. It is possible to retrieve primer sequences as well as a graphical representation of a chromosome map that shows the microsatellites (in Macintosh PICT format suitable for incorporation into all Mac drawing programs). To receive a blank E-mail query form and instructions for its use, send an E-mail message with the single word help to genome_database@genome.wi.mit.edu. Data can also be retrieved by anonymous ftp to genome.wi.mit.edu. For more information, contact Lincoln Stein at the Whitehead Institute [Email address: lstein@genome.wi.mit.edu].

B.1.2 Humans and other organisms

A central Gopher Hole for human genetic information is maintained at Johns Hopkins University. It can be reached by tracing a path through Maryland to Johns Hopkins University to "Computational Biology" or by means of the Jackson Laboratory Gopher hole described above. Once there, you should work your way down through the "Genome Project" to the "Genome Data Base (GDB) folder." Once you have reached this location, it becomes possible to search various databases of human genetic information including the Genome Database (GDB) itself, which is a compendium of human mapping data, other genetic information, and citations. At this Gopher Hole, it is also possible to search the electronic version of Victor McKusick’s Mendelian Inheritance of Man (called OMIM) which contains comprehensive records on all defined human loci. For further information, the GDB staff can be reached at the E-mail address [Help@gdb.org] or the mailing address [GDB User Support, Genome Data Base, Johns Hopkins University, 2024 E. Monument Street, Baltimore, MD 21205-2100.]

Other Gopher Holes dedicated to the human genome project are maintained at the Genethon Center in France (point to [Gopher.genethon.fr 70]), and the UK Mapping Project (point to [Menu.crc.ac.uk 70]). A database of Drosophila mapping information is maintained at Indiana University (point to [fly.bio.indiana.edu]) and the yeast database is maintained at Stanford University (point to genome.stanford.edu]). Mapping information on C. elegans is maintained at the MRC in Cambidge, England; contact Richard Durbin for further information (E-mail address is [rd@mac-lib.cam.ac.uk]).

B.2 Mouse Genetics Community Bulletin Board

An electronic Bulletin Board (called MGI-LIST) has been set up at the Jackson Laboratory with E-mail addresses of mouse geneticists throughout the world. The Mouse Genome Informatics Group at the JAX will use this Bulletin Board to make announcements of new software and data sets for the Encyclopedia of the Mouse Genome described below. In addition, all subscribers will be able to broadcast to, and receive messages of interest from, all other linked-up members of the community. If you would like to subscribe to this Bulletin Board, send a message that reads "subscribe mgi-list <your name>" to listserver@informatics.jax.org. For further information or help, send an E-mail message to mgi-help@informatics.jax.org.

B.3 Computer Programs

B.3.1 Linkage analysis

Numerous computer programs are available for the determination of linkage relationships among loci typed in large numbers of individuals. Most of these were developed for the analysis of complex human pedigrees with algorithms that incorporate Maximum Likelihood Estimation statistics (Elston and Stewart, 1971) and provide LOD score (Morton, 1955) output. Although these pedigree-based programs can be used for the analysis of mouse linkage data, they are usually less than optimal for this task. First, it is often not possible to derive unequivocal haplotype information for all individuals studied in a human pedigree, and as such, pedigree-based programs are not oriented toward this type of analysis which is so common in mouse linkage studies (see figure 9.15). Second, for most studies, mouse geneticists will not need to avail themselves of the sophisticated Maximum likelihood/LOD score estimation tools required for human pedigree analysis. Two linkage programs developed specifically for mouse linkage studies — Map Manager and GENE-LINK — are described below. In addition, the most useful multi-locus pedigree-based program — Mapmaker — is also described. Other pedigree-based linkage analysis packages are listed and reviewed by Bryant (1991).

Map Manager is a Macintosh program that can be used for the analysis of linkage data obtained from a backcross, F1 X F1 intercross, or from a group of recombinant inbred strains (Manly, 1993). It was written by Dr. Kenneth F. Manly and is made available without charge from the author. Map Manager uses a standard Macintosh format and is both extremely versatile and extremely easy to use. It allows easy storage, display, and retrieval of information from mapping experiments. The strain distribution patterns (SDPs) obtained with newly typed loci can be evaluated rapidly to determine likely map positions relative to other loci in the database. The program is distributed with a large database of previously published RI strain distribution patterns which greatly facilitates the RI analysis of new loci. Map Manager has many other sophisticated features including a statistical evaluation package and various output options, as well as a means for importing and exporting data to and from spreadsheets or Mapmaker files (see below). It can be obtained from the author on disk or by anonymous ftp or gopher from several sites including {mcbio.med.buffalo.edu}, {hobbes.jax.org}, {ftp.bio.indiana.edu}, and {ftp.embl-heidelberg.de}. For further information, Dr. Manly can be contacted at the E-mail address [Kmanly@mcbio.med.buffalo.edu] or at Roswell Park Cancer Institute, Buffalo, New York 14263.

GENE-LINK is a DOS program that can be used for the analysis of linkage data from a backcross (Montagutelli, 1990). It was written by Dr. Xavier Montagutelli and has been made available to investigators without cost. It will provide best-fit map positions for entered strain distribution patterns. For further information, Dr. Montagutelli can be contacted at Institut Pasteur de Paris, 25 Rue du Docteur Roux, 75724 Paris, Cedex 15, France.

Mapmaker and Mapmaker/QTL are a pair of pedigree-based programs written by Dr. Eric Lander and his colleagues for constructing linkage maps from raw genotyping and phenotyping data recovered from large numbers of loci (Lander et al., 1987). These programs use a highly efficient algorithm for "likelihood of linkage" computations. They can be run on UNIX-based workstations or VAX minicomputers running under the VMS operating system. The programs and a manual are available from the author for licensing to academic researchers. Mapmaker is most useful to mouse geneticists for the analysis of linkage data obtained from an F1 x F1 intercross. Mapmaker/QTL can be used for the analysis of quantitative traits. For further information, contact Dr. Eric Lander, Whitehead Institute, 9 Cambridge Center, Cambridge, MA 02142.

B.3.2 Mouse colony management and genetic databases

MacMice and Mendel's Lab are Macintosh-based programs that can be used for keeping track of a breeding mouse colony with records for animals, litters, and samples derived from them. The programs are described more fully in chapter 3 (Silver, 1986; Silver, 1993b). They were written by the author of this book and can be licensed for use from Mendel's Software. Click on the following site: for further information and ordering details, or send a FAX to Mendel's Software at 609-924-4382.

Appendix C

Source Materials for Further Reading

C.1 Selected monographs and books

C.1.1 General

C.1.2 Genetic information

C.1.3 Experimental embryology

C.1.4 Others topics

C.2 Selected Journals and Serials of Particular Interest

C.2.1 Original articles

C.2.2 Reviews

 

C.1 Selected monographs and books

C.1.1 General

Green, M. C. & Witham, B. A. (1991). Handbook on Genetically Standardized JAX Mice, Fourth edition, The Jackson Laboratory, Bar Harbor. [Provided free upon request.]

Foster, H. L., Small, J. D. & Fox, J. G., eds. (1982). The Mouse in Biomedical Research, Diseases. Academic Press, New York. [A series of four volumes with comprehensive coverage of all aspects of mouse biology and breeding]

Altman, P. L. & Katz, D. (1979). Inbred and Genetically Defined Strains of Laboratory Animals: Part I, Mouse and Rat., Biological Handbooks , 3rd edition, Fed. Amer. Soc. Exp. Biol., Bethesda. [Tables of information for different strains of mice.]

Klein, J. (1975). Biology of the Mouse Histocompatibility-2 Complex, Springer-Verlag, New York. [The title is misleadingly narrow; a general discussion of mouse genetics.]

Crispens, C. G. (1975). Handbook on the Laboratory Mouse, C.C. Thomas, Springfield, Ill. [Information on strains not commonly used today.]

Green, E. L., ed. (1966). Biology of the Laboratory Mouse, 2nd edition, McGraw-Hill, New York. [Reprinted by Dover (New York) in 1968 and 1975; Comprehensive for its time.]

C.1.2 Genetic information

Mammalian Genome: Special Issue published annually by Springer-Verlag, New York, with reports from the mouse chromosome mapping committees.

Lyon, M. F. & Searle, A. G., eds. (1989). Genetic Variants and Strains of the Laboratory Mouse., 2nd edition, Oxford University Press, Oxford. [The "Bible" for mouse geneticists: description and references for all known mutant loci along with other genetic information. Look for newer edition.]

Silvers, W. K. (1979). The Coat Colors of the Mouse, Springer-Verlag, New York. [Excellent coverage of this particular subject. Sadly, now out-of-print.]

C.1.3 Experimental embryology

Hogan, B., Beddington, R., Costantini, F. & Lacy, E. (1994). Manipulating the Mouse Embryo: A laboratory Manual, Second Edition, Cold Spring Harbor Laboratory Press, Cold Spring Harbor. [The bible for embryological manipulations.]

Wassarman, P. M. & DePamphilis, M. L., eds. (1993). Guide to Techniques in Mouse Development, Methods in Enzymology , 225, Academic Press, San Diego. [Comprehensive coverage of techniques used for developmental analysis.]

Kaufman, M. H. (1992). An Atlas of Mouse Development, Academic Press, San Diego. [Photomicrographs of embyo sections.]

Rugh, R. (1968). The Mouse: Its Reproduction and Development, Oxford University Press, New York. [Source of information.]

C.1.4 Others topics

Berry, R. J. & Corti, M., eds. (1990). Biological Journal of the Linnean Society, volume 41, Academic Press, London. [Proceedings from a conference on the evolution, population biology, and systematics of the mouse.]

Potter, M., Nadeau, J. H. & Cancro, M. P., eds. (1986). The Wild Mouse in Immunology, Curr. Topics Micro. Immunol. , volume 127, Springer-Verlag, New York. [Proceedings from a conference on the evolution and systematics of the mouse.]

Klein, J. (1986). Natural History of the Major Histocompatibility Complex, John Wiley & Sons, New York. [Comprehensive discussion of various topics in mouse genetics, evolution, and population biology.]

Green, E. L. (1981). Genetics and Probability in Animal Breeding Experiments, Oxford University Press, New York. [General presentation of the statistical principles of genetic analysis.]

Morse, H. C., ed. (1978). Origins of Inbred Mice, , Academic Press, New York. [Interesting proceedings on historical aspects.]

Simmons, M. L. & Brick, J. O. (1970). The Laboratory Mouse: Selection and Management, Prentice-Hall, Englewood Cliffs, NJ.

Cook, M. J. (1965). The Anatomy of the Laboratory Mouse, Academic Press, New York. [An atlas of anatomical drawings].

C.2 Selected Journals and Serials of Particular Interest

C.2.1 Original articles

Cell

Cytogenetics and Cell Genetics

Genes and Development

Genetical Research

Genetics

Genomics

Immunogenetics

Journal of Heredity

Mammalian Genome

Mouse Genome (formerly Mouse Newsletter)

Nature

Nature Genetics

Proceedings of the National Academy of Science USA

Science

C.2.2 Reviews

Annual Review of Genetics

Current Opinion in Genetics and Development

Trends in Genetics

Appendix D

Statistics

D.1 Confidence limits and median estimates of linkage distance

D.1.1 The special case of complete concordance

D.1.2 The general case of one or more recombinants

D.1.3 A C program for the calculation of linkage distance estimates and confidence intervals

D.2 Quantitative differences in expression between two strains

D.1 Confidence Limits and Median Estimates of Linkage Distance

D.1.1 The special case of complete concordance

To illustrate the statistical approach used to estimate confidence limits on experimentally-determined values for linkage distances, it is useful to first consider the special case where two linked loci show complete concordance or no recombination (symbolized asR = 0) in their allelic segregations among a set of N samples derived either from recombinant inbred strains or from the offspring of a backcross. Let us define the true recombination fractionQ — as the experimental fraction of samples expected to be discordant (or recombinant) when N approaches infinity. Then the probability of recombination in any one sample is simply Q, and the probability of non-recombination, or concordance, is simply (1 - Q). As long as multiple events are completely independent of each other, one can calculate the probability that all of them will occur by multiplying together the individual probabilities associated with each event. Thus, if the probability of concordance in one sample is (1 — Q), then the probability of concordance in N samples is: .

In most experimental situations, the known and unknown variables are reversed in that one begins by determining the number of discordant (or recombinant) samples i that occur within a total set of N as a means to estimate the unknown true recombination fraction Q. When no discordant samples are observed, the probability term just derived can be used with the substitution of the random variable q in place of Q, to provide a continuous probability density function indicative of the relative likelihoods for different values of Q between 0.0 (complete linkage) and 0.5 (no linkage).

? ?(D.1)

This equation reads "the probability that the true recombination fraction Q is equal to a particular value q is the function of q given as the last term in the equation". For both RI data and backcross data, Q is related directly to linkage distance in centimorgans d. In the case of backcross data, and for values of Q less than 0.20 (see section 7.2.2.3), recombination fractions are converted into centimorgan estimates through simple multiplication:

?d = 100Q. ?(D.2)

In the case of RI data, this conversion is combined with the Haldane-Waddington equation (9.8) to yield:

? ?(D.3)

An example of the probability density function associated with the experimental observation of complete concordance among 50 backcross samples is shown in figure D.1. Each value of N will define a different function, but in all cases, the curve will look the same with only the steepness of the fall-off increasing as N increases. In all cases, the "maximum likelihood estimate" for the true recombination fraction — defined as the value of Q associated with the highest probability — will be zero. But, since this maximum likelihood value is located at one end of the probability curve, it does not provide a useful estimate for the likely linkage distance. A better estimate would be the value of Q which defines the midpoint below which and above which the true recombination fraction value is likely to lie with equal probability; this is the definition of the median recombination fraction estimate . In mathematical terms, the value of is defined at the line which equally divides the area of the complete probability density given by equation D.1 (see figure D.1).

Confidence limits are also defined by circumscribed portions of the entire probability density; the portion that lies outside a confidence interval is called a. For example, in the case of a 95% confidence inerval, a = (1 - 0.95) = 0.05. It is standard practice to assign equal portions of a to the two "tails" of the probability density located before and after the central confidence interval. Thus, the lower confidence limit is defined as the value of q bordering the initial a/2 fraction of the area under the entire probability curve. The upper confidence limit is defined as the value of q that borders the ultimate a/2 fraction of the area under the entire probability curve; this is equivalent to saying that a ‘(1 — a/2)’ fraction of area lies ahead of the upper confidence limit.

In mathematical terms, the area beneath the entire probability density curve is equal to the definite integral of equation D.1 over the range of legitimate values for q between 0.0 and 0.5. To determine the fraction of the probability density that lies in the region between Q = 0 and any arbitrary Q = x, it is necessary to integrate over the probability density function (equation D.1) between these two values, and divide the result by the total area covered by the probability density. This provides the probability that the true recombination fraction is less than or equal to x.

? ?(D.4)?

By standard methods of calculus, equation D.4 can be reduced analytically to the form:

? ?(D.5)

And this equation can be reformulated to yield x as a function of .

? ?(D.6)?

By solving equation D.6 for different values of , one can obtain critical values of x that define the median estimate of the recombination fraction from , lower confidence limits from , and upper confidence limits from . Once a solution for x has been obtained, it can be converted into a linkage distance value with either equation D.2 for backcross data or equation D.3 for RI strain data. Solutions to equation D.6 over a range of N RI strains and backcross animals are shown in figures 9.8, 9.16, and 9.17.

D.1.2 The general case of one or more recombinants

The statistical approach described above can be generalized to any case of i discordant (or recombinant) samples observed among a total of N RI strains or backcross animals that have been typed for two loci. As in the special case above, one can arrive at a probability for the occurrence of multiple events by multiplying together the individual probabilities for each event. In the general case, there will be i events of discordance, each with an individual probability equal to the true recombination fraction Q, and (Ni) events of concordance, each with an individual probability of (1 — Q). These terms are multiplied together along with a "binomial coefficient" that counts the the permutations in which the two types of events can appear to produce the "binomial formula":

? ?(D.7)

When the true recombination fraction is known, the binomial formula can be used to provide the probability that i events of discordance will be observed in any set of N samples. But once again, the situation encountered by geneticists is usually the reverse one in which i and N are discrete values determined by the experiment and the true recombination fraction Q is unknown. In this case, one can substitute the random variable q in place of Q in equation D.7 to generate a probability density function that provides relative likelihoods for different values of Q between 0.0 (complete linkage) and 0.5 (no linkage). In this use of the binomial formula, the factorial fraction (known as the binomial coefficient) remains constant for all values of q and can be eliminated since the purpose of the function is to provide relative probabilities only:

? ?(D.8)

An example of the probability density function associated with the experimental observation of one discordant RI strain among a total of 26 samples is shown in figure D.2. As one can easily see, the distribution is highly skewed toward higher recombination fractions. Each discrete pair of values i and N will define a different function. When both i and N are large, the density function will approximate a normal distribution. However, with the results typically obtained in contemporary mouse linkage studies, the density function is likely to be significantly skewed as shown in figure D.2 and as such, it is usually not possible to take advantage of the simplified statistical tools developed specially for use with the normal distribution.

A median estimate of linkage distance as well as lower and upper confidence limits can be obtained in the same manner described in the special case of no recombination described above. This can be accomplished by substituting equation D.8 in place of the two occurrences of equation D.1 within equation D.4:

? ?(D.9)

The general form of the integral in this equation cannot be solved analytically but a short computer program can be used to estimate solutions and provide critical values of x for defined probability values. The computer program has been written to generate minimum and maximum values in terms of centimorgan distances for discrete experimentally-determined values of i and N from either backcross or RI data. The program was used to generate the values shown in Tables D.1, D.2, D.3, D.4, D.5, and D.6 for 68% and 95% confidence intervals, but it is possible to generate confidence limits for any other integer percentile confidence interval. The program will also calculate maximum likelihood and median estimates of linkage distance. It is listed below as a self-contained unit that should be ready for compiling with any standard C compiler on any computer.

 

D.1.3 A C program for the calculation of linkage distance estimates and confidence intervals

/*** A C program for the calculation of linkage distance estimates and confidence intervals ***/

#include <stdio.h>

double ?Pin(double r,int i,int N); double pow(double x, double y); double convert(double r);

static?int crosstype;

main()

{?FILE ?*fopen(), *file;

?int ?i=1, istart=1, ifin=50, iinc=1, N=100, P;

?char?input;

?double?Pin(), dmin, dmax, r,rtop, dmean, smean, Nrmlize=0.0, Sum=0.0, convert(), min, max;

?while(1){

??printf("Enter the type of cross:1 for backcross,2 for RI analysis,or 3 to quit:");

??scanf("%d",&crosstype);

??if(crosstype != 2 && crosstype != 1) exit(0);

??printf("Enter the confidence level as an integer number(e.g. 95 for 95%%):");

??scanf("%d", &P);

??min = (1-((double)P/100.0))/2; max = 1- min;

??printf("Enter with comma delimiters->i-start,i-end,i-increment,and N,then return\n:>");

??scanf ("%d,%d,%d,%d", &istart,&ifin,&iinc,&N);

??printf(" i , dist / medn , min. / max. (values in cM assuming complete interference)\n");

??for ( i = istart; i <= ifin ; i += iinc){

???for ( r = .0001, Nrmlize = 0 ; r <.5 ; r += .0001)

????Nrmlize += Pin(r,i,N);

???for ( r = .0001, Sum =0; Sum < min && r< .5; r += .0001)

????Sum += Pin(r,i,N)/Nrmlize;

???dmin = convert(r);

???for (; Sum < .5 && r< .5; r += .0001)

????Sum += Pin(r,i,N)/Nrmlize;

???dmean = convert(r);

???for (; Sum < max && r< .5 ; r += .0001)

????Sum += Pin(r,i,N)/Nrmlize;

???dmax = convert(r);

???smean = convert((double)i/N);

???printf("%3d, %4.1f / %4.1f , %4.1f / %4.1f\n",i,smean,dmean,dmin,dmax);}

?}}

double convert(double r)

{?double rmean;?int x=0;

?if(crosstype == 1)?return(100*r);

?if(crosstype == 2)?return( r*100/(4 - 6*r) );}

double Pin(double r ,int i ,int N)

{?double?pow();

?return ((pow(r,i))*(pow(1-r,N-i)));}

/************************ END OF PROGRAM ***********************/

 

D.2 Quantitative differences in expression between two strains

How does one determine whether two populations of animals defined by different inbred strains are showing a significant difference in the expression of a trait? The answer is with a test statistic known as the "t-test" or "Student’s t-test". To apply this test, one needs to use a pair of only three values derived from an analysis of the expression of the trait in sets of animals from each inbred strain. First is the number of animals examined in each inbred set (N1 and N2). Second is the mean level of expression for each set (m1 and m2) calculated as:

? ?(D.10)

where xi refers to the expression value obtained for the ith sample in the set. Third is the variance of each set of animals ( and ) calculated as:

? ?(D.11)

With values for the variance of each sample set and the size of each set, one can calculate a combined parameter refered to as the "pooled variance":

? ?(D.12)

Finally, one can use the value obtained for the pooled variance together with the samples sizes and sample means to obtain a "t value":

? ?(D.13)

One final combined parameter is required to convert the t value into a level of significance — the number of degrees of freedom df.

? ?(D.14)

With values for t and df, one can obtain a P value from a table of critical values for the t distribution found in Table D.7.

Appendix E

Glossary of Terms

Allele An alternate form of a gene or locus. A locus can have many different alleles which may differ from each other by as little as a single base or by the complete absence of a sequence.
Allele-Specific Oligonucleotide (ASO) An oligonucleotide designed to hybridize only to one of two or more alternative alleles at a locus. An ASO is usually designed around a variant nucleotide located at or near its center (see chapter 8).
Anchor locus A well-mapped locus that is chosen as a marker to "anchor" a particular genomic region to a framework map that is being constructed in a linkage study with a new cross (see chapter 9).
Anonymous locus An isolated DNA region with no known function but with at least two allelic states that can be followed through some form of DNA analysis in mapping studies.
ASO See Allele-Specific Oligonucleotide.
B1 repeat The most prominent SINE class of highly dispersed repetitive elements in the genome with a copy number of ~150,000 (see chapter 5).
B2 repeat The second most prominent SINE class of highly dispersed repetitive elements in the genome with a copy number of ~90,000 (see chapter 5).
B6 An abbreviation for the name of the most commonly used strain of mice — C57BL/6.
Bayesian analysis A statistical approach that takes prior information into account in the determination of probabilites. The Bayesian approach yields an equation that must be used to convert P values obtained from c2 analysis of recombination data into actual probabilities of linkage between two loci (chapter 9).
Backcross A cross between one animal type that is heterozygous for alleles obtained from two parental strains and a second animal type from one of those parental strains. The term is often used by itself to describe the two generation breeding protocol of an outcross followed by a backcross used frequently in linkage analysis (see chapters 3 and 9).
CA-repeat The most prominent class of microsatellites found in mammalian genomes (see chapter 8).
castaneus The shortened form of M. m. castaneus, a subspecies within the M. musculus group that can be combined with a traditional inbred strain for linkage analysis (see chapters 2 and 9).
centimorgan (cM) The metric used to describe linkage distances. A centimorgan is the distance between two genes that will recombine with a frequency of exactly one percent. This term is named afer Thomas Hunt Morgan, who first conceptualized linkage while working with Drosophila.
Chi-squared A statistical test used most often by geneticists to ascertain whether experimental data provide significant evidence for linkage between two loci (see chapter 9).
Chimera An individual mouse, or other mammal, that is derived from the fusion of two or more preimplantation embryos or an embryo and ES cells (see chapter 6).
Chromosome bands Alternative light and dark-staining regions within chromosomes that are visualized by light microscopy (see chapter 5).
cM See centimorgan.
Codominance Defined for pairs of alleles. The situation in which an animal heterozygous for two alleles (A1 and A2 at the A locus) expresses both of the phenotypes observed in the two corresponding homozygotes. Thus, the heterozygote (A1/A2) and both homozygotes (A1/A1 and A2/A2) are all distinguishable from each other and A1 and A2 would be considered to be "codominant". This term has also been coopted to describe DNA markers defined by alternative visible allelic forms such as different sized restriction fragments or PCR products.
Coisogenic A variant strain of mice that differs from an established inbred strain by mutation at only a single locus (see chapter 3).
Commensal Pertaining to populations of house mice that depend on human-built habitats and/or food production for survival (see chapter 2).
Concordance For two or more loci or traits typed in offspring from a backcross or RI strain, the presence of alleles (or expression of a trait) derived from the same parental chromosome (see chapter 9).
Congenic A variant strain of mice that is formed by backcrossing to an inbred parental strain for ten or more generations while maintaining heterozygosity at a selected locus (see chapter 3).
Conplastic A variation on the congenic approach in which the mitochondrial genome from one strain is transferred onto a different genetic background (see chapter 3).
Consomic A variation on the congenic approach in which an entire chromosome from one strain, usually the Y, is transferred onto a different genetic background (see chapter 3).
Contig A set of contiguous overlapping genomic clones that together span a larger region of the genome than that covered by any one clone (see chapters 7 and 10).
CpG island A genomic region of one or a few kilobases in length that contains a high density of CpG dinucleotides. CpG islands are associated with the 5’-ends of genes (see chapter 10; previously called an HTF island).
Cross Refers to one or more mating units set up with males and females that each have a designated genotype chosen to carry out a particular genetic analysis (see chapter 3).
Crossover product A chromosome homolog that was formed through the recombination of alleles at different loci present on opposite homologs in one of the parents of the animal in which it is observed (see chapter 7).
Deme A breeding group unit. In natural populations of mice, a deme usually consists of one breeding male with a harem of up to 8 females (see chapter 2).
Differential segment In the genome of a congenic mouse, the region of chromosome surrounding the selected locus that is derived together with it from the donor genome (see chapter 3).
Discordance The opposite of concordance. Inheritance of only one of two alleles or traits associated with a particular parental chromosome.
Disjunction The normal process by which the two homologs of each chromosome in a meiotic cell separate and move to different gametes (see chapter 5).
Distal A relative term meaning closer to the telomere; the opposite of proximal.
DNA marker A cloned chromosomal locus with allelic variation that can be followed directly by a DNA-based assay such as Southern blotting or PCR (see chapter 8).
domesticus Short form of M. m. domesticus, a subspecies within the M. musculus group that resides in Western Europe, Africa, and throughout the New World. It is the primary component of the traditional inbred strains (see chapter 2).
Dominant A relative term describing the relationship of one allele to a second at the same locus when an animal heterozygous for these alleles expresses the same phenotype as an animal homozygous for the first allele. The second allele of the pair is considered recessive.
ES cells Embryonic stem cells. Cultured cells derived from the pluripotent inner cell mass of blastocyst stage embryos. Used for gene targeting by homologous recombination (see chapter 6).
Expressivity Pertaining to observed quantitative differences in the expression of a phenotype among individuals that have the same mutant genotype. When quantitative differences are observed, a phenotype is said to show "variable expressivity" which can be caused by environmental factors, modifier genes or chance.
F1 The first filial generation; the offspring of an outcross between two inbred strains (see chapter 3).
Feral pertaining to wild populations of animals derived from commensal ancestors; house mice that live apart from, and independent of, humans (see chapter 2).
Filial generation Pertaining to a particular generation in a sequence of brother-sister matings that can be carried out to form an inbred strain. The first filial generation, symbolized as F1, refers to the offspring of a cross between mice having non-identical genomes. When F1 siblings are crossed to each other, their offspring are considered to be members of the second filial generation or F2, with subsequent generations of brother-sister matings numbered with integer increments (see chapter 3).
FISH Fluorescent In situ Hybridization. An enhanced from of in situ hybridization with high resolution and sensitivity (see chapter 10).
Genetic drift Refers to the constant tendency of genes to evolve even in the absence of selective forces. Genetic drift is fueled by spontaneous neutral mutations that disappear or become fixed in a population at random. Inbred lines separated from a common ancestral pair can drift rapidly apart from each other.
Genome The total genetic information present within a single cell nucleus of an animal. The haploid genome content of the mouse is 3 x109 bp.
Genotype For any one animal, the set of alleles present at one or more loci under investigation. At any one autosomal locus, a genotype will be either homozygous (with two identical alleles) or heterozygous (with two different alleles).
Giemsa A stain and associated protocol used to accentuate visually the difference between bands and interbands on metaphase chromosomes (see chapter 5).
Haplotype Pertaining to a particular set of alleles at linked loci (or nucleotide changes within a gene) that are found together on a single homolog. In linkage studies with backcross offspring and RI strains, the haplotypes associated with each sample provide a means for determining the order of loci (see chapter 9).
Heterozygote An animal with two distinguishable alleles at a particular locus under analysis. In this case, the locus is considered to be heterozygous.
Histocompatible Pertaining to a genetic state in which cells from two animals can be cross-transplanted without immunological rejection. The opposite of histoincompatible. Histocompatibility is controlled predominantly by genes in the Major Histocompatiblity Complex or MHC (see chapter 3).
Homolog This term is used by geneticists in two different senses: (1) One member of a chromosome pair in diploid organisms, and (2) A gene from one species, for example the mouse, that has a common origin and functions the same as a gene from another species, for example humans, Drosophila, or yeast.
Homozygote An animal with two identical alleles at a particular locus under analysis. In this case, the locus is considered to be homozygous.
Hotspot, recombinational A localized region of chromosome, usually less than a few kilobases in length, that participates in crossover events at a very high rate relative to neighboring regions of chromosome (see chapter 7).
House mouse An animal that is a member of the species M. musculus.
HTF island See CpG island.
Hybrid sterility Pertaining to the sterility of animals produced from matings between members of two different species, such as M. musculus and M. spretus. In this case, and in general, only the male hybrids are sterile while the females are fertile (see chapter 2).
Hybrid zone A narrow geographical line that separates the natural ranges of two distinct animal populations. The best-characterized house mouse hybrid zone occurs in Central Europe and separates M. m. domesticus to the west and M. m. musculus to the east (see chapter 2).
Imprinting, Genomic The situation in which the expression of a gene varies depending on its parental origin (see chapter 5). Only a small subset of genes in the mammalian genome are imprinted.
In situ hybridization A technique for mapping cloned DNA sequences by hybridization directly to metaphase chromosomes and analysis by microscopy (see chapter 10).
Inbred Animals that result from the process of at least twenty sequential generations of brother-sister matings. This process is called inbreeding (see chapter 3).
Incross A cross between two animals that have the same homozygous genotype at designated loci; for example, between members of the same inbred strain (see chapter 3).
Intercross A cross between two animals that have the same heterozygous genotype at designated loci; for example, between sibling F1 hybrids that were derived from an outcross between two inbred strains (see chapter 3).
Interference The suppression of crossing over that occurs in the extended chromosomal vicinity of an initial crossover event. Interference is responsible for a severe reduction in the expected frequency of double crossover events in ten to twenty centimorgan lengths of the genome (see chapter 7).
Interspecific cross A cross between mice from two different species, most often M. musculus (represented by a traditional laboratory strain) and M. spretus for the purpose of linkage analysis. The interspecific cross is carried out to take advantage of the high level of polymorphism between the two parents (see chapter 9).
Intersubspecific cross A cross between two subspecies (see chapter 9). In the case of mouse genetics, this refers most often to a cross between a traditional inbred strain that is predominantly M. m. domesticus and one of the other subspecies in the M. musculus complex, usually M. m. musculus or M. m. castaneus or a combination of both (within the faux species M. m. molossinus).
IRS PCR Interspersed Repetitive Sequence PCR. A method for amplifying species-specific sequences from a complex hybrid genome (see chapter 8).
Karyotype The number of chromosomes present in a given genome and the form that they assume (including banding patterns) when they condense (see chapter 5). A karyotype is defined entirely by microscopic observation.
Library, genomic A sufficient number of genomic clones such that any sequence of interest is very likely to be present in at least one member of the set. If the library is random, the actual set of original clones must contain a cumulative length of DNA that is equal to multiple "genomic equivalents."
Linkage Pertaining to the situation where two loci are close enough to each on the same chromosome such that recombination between them is reduced to a level significantly less than 50%.
Linkage group A set of loci in which all members are linked either directly or indirectly to all other member of the set. Essentially equivalent to the genetic information associated with any single chromosome.
Locus Any genomic site, whether functional or not, that can be mapped through formal genetic analysis.
Meiosis The process by which diploid germ cell precursors segregate their chromosomes into haploid nuclei within eggs and sperm.
Meiotic product An individual haploid genome within an egg or sperm cell. Meiotic products are usually observed and analyzed within the context of diploid offspring.
Microdissection A method for dissecting and cloning from defined subchromosomal regions by microscopic examination and manipulation (see chapter 8).
Microsatellite A very short unit sequence of DNA (2 to 4 bp) that is repeated multiple times in tandem. Microsatellites (also called Simple Sequence Repeats or SSRs) are highly polymorphic and make ideal markers for linkage analysis (see chapter 8). A polymorphism at a microsatellite locus is also referred to as a Simple Sequence Length Polymorphism (SSLP).
Minisatellite A highly polymorphic type of locus containing tandemly repeated sequences having a unit length of 10 to 40 bp. Minisatellite polymorphisms can be assayed by RFLP analysis or by PCR (see chapter 8). Also referred to as Variable Number of Tandem Repeat (VNTR) loci.
Multifactorial A trait controlled by at least two factors, which may be genetic or environmental (see chapter 9); polygenic traits represent a subset of multifactorial traits.
Mus the name of the genus that contains all house mice (M. musculus) and other closely related species.
musculus Abbreviated form of M. musculus, the species that is synonymous with the house mouse (see chapter 2).
Mutant allele This term is defined differently by formal geneticists and population biologists. The formal genetic definition is an allele that exerts a deleterious effect on phenotype. The population definition is an allele that is present at a frequency of less than 1% in a natural population; according to this definition, a mutant allele in one population may be considered non-mutant (wild-type) in another population.
Mutation An allele present in a progeny that is not present in the genome of either its parents.
N2, N3, N4 etc. Used to describe the generation of backcrossing and the offspring that derive from it. The N2 generation describes offspring from the initial cross between an F1 hybrid and one of the parental strains. Each following backcross generation is numbered in sequence (see chapter 3).
Outcross A cross between genetically unrelated animals.
Parental An inbred strain of animals that is used in the initial cross of a multi-generational breeding protocol; the meiotic products and offspring that retain the same set of designated alleles as one of the parental strains.
Penetrance Pertaining to the failure of some animals with a mutant genotype to express the associated mutant phenotype. In any case where less than 100% of genotypically mutant animals are phenotypically mutant, the phenotype is said to be "incompletely penetrant." Incomplete penetrance is usually a matter of chance or modifiers in the genetic background.
PFGE Pulsed Field Gel Electrophoresis. A technique for separating very large DNA molecules from each other (see chapter 10).
Phenotype The physical manifestation of a genotype within an animal. A mutant phenotype is caused by a mutant genotype and is manifested as an alteration within an animal that distinguishes it from the wild-type.
Phylogenetic tree A diagram showing the postulated evolutionary relationships that exist among related species in terms of their divergence from a series of common ancestors at specific points in time.
Polygenic Pertaining to a phenotype that results from interactions among the products of two or more genes with alternative alleles (see chapter 9).
Polymorphic A term formulated by population geneticists to describe loci at which there are two or more alleles that are each present at a frequency of at least one percent in a population of animals. The term has been co-opted for use in transmission genetics to describe any locus at which at least two alleles are available for use in breeding studies, irrespective of their actual frequencies in natural populations.
Proximal A relative term meaning closer to the centromere; the opposite of distal.
Quantitative trait A phenotype that can vary in a quantitative manner when measured among different individuals (see chapter 9). The variation in expression can be due to combinations of genetic and environmental factors, as well as chance. Quantitative traits are often controlled by the cumulative action of alleles at multiple loci.
Recessive A relative term describing the relationship of one allele to a second at the same locus when an animal heterozygous for these alleles expresses the same phenotype as an animal homozygous for the second allele. The second allele of the pair is considered dominant.
Recombinant The result of a crossover in a doubly heterozygous parent such that alleles at two loci that were present on opposite homologs are brought together on the same homolog. The term is used to describe the chromosome as well as the animal in which it is present.
Recombinant congenic strain A variation on recombinant inbred strains in which the initial outcross is followed by several generations of backcrossing prior to inbreeding (see chapter 3).
Recombinant inbred (RI) strain A special type of inbred strain formed from an initial outcross between two well-characterized inbred strains followed by at least twenty generations of inbreeding (see chapter 9).
Restriction Fragment Length Polymorphism (RFLP) A DNA variation that affects the distance between restriction sites (most often by a nucleotide change that creates or eliminates a site) within or flanking a DNA fragment recognized by a cloned probe (see chapter 8). RFLPs are detected upon Southern blot hybridization. The term RFLP is commonly used even in situations where the DNA variation may not represent a true polymorphism in the population-based definition of this term.
Restriction Fragment Length Variant (RFLV) A more accurate term to use in place of Restriction Fragment Length Polymorphism in those cases where the frequency of the variant in natural populations is not known.
Retroposon An inserted genomic element that originated from the reverse transcribed mRNA produced from another region of the genome (see chapter 5).
RFLP See Restriction Fragment Length Polymorphism.
RFLV See Restriction Fragment Length Variant
RI strain See Recombinant Inbred strain
Robertsonian translocation A fusion between the centromeres of two acrocentric chromosomes to produce a single metacentric chromosome (see chapter 5).
Satellite DNA This term was used originally to describe a discrete fraction of DNA visible in a CsCl2 density gradient as a "satellite" to the main DNA band. The term now refers to all simple sequence DNA having a centromeric location, whether distinguishable on density gradients or not (see chapter 5).
SDP See Strain Distribution Pattern.
Simple Sequence Repeat (SSR) See microsatellite.
SINE Short INterspersed Element. Families of selfish DNA elements that are a few hundred basepairs in size and dispersed throughout the genome (see chapter 5).
spretus Abbreviated form of M. spretus, a species commonly used in interspecific matings for the generation of linkage maps (see chapters 2 and 9).
SSCP Single Strand Conformation Polymorphism. A gel-based means for detecting single nucleotide changes within allelic PCR products that have been denatured and gel-fractionated as single strands.
SSLP Simple Sequence Length Polymorphism; see microsatellite.
SSR Simple Sequence Repeat; see microsatellite.
Strain Refers to a group of mice that are bred within a closed colony in order to maintain certain defining characteristics. Strains can be inbred or non-inbred (see chapter 3).
Strain Distribution Pattern (SDP) The distribution of two segregating alleles at a single locus across a group of animal samples used for analysis in a linkage study (see chapter 9). Used in the context of backcross data and data obtained from RI strains.
Sympatric Closely related species that have overlapping ranges in nature but do not interbreed. In different parts of its range, M. musculus is sympatric with M. macedonicus, M. spicilegus, and M. spretus (see chapter 2).
Syngenic Literally "of the same genotype." Used most frequently by immunologists to describe interactions between cells from the same inbred strain.
Syntenic Two loci known to be in the same linkage group. Conserved synteny refers to the situation where two linked loci in one species (such as the mouse) have homologs that are also linked in another species (such as humans).
Targeting, Gene A technology that allows an investigator to direct mutations to a specific locus in the mouse genome (see chapter 6). Also called targeted mutagenesis.
Transgene A fragment of foreign DNA that has been incorporated into the genome through the manipulation of preimplantation embryos (see chapter 6).
Transgenic Pertaining to an animal or locus that contains a transgene (see chapter 6).
Translocation Pertaining to a novel chromosome formed by breakage and reunion of DNA molecules into a non-wild-type configuration (see chapter 5).
Unequal crossover A crossover event that occurs between non-allelic sites. Can lead to the duplication of sequences on one homolog and the deletion of sequences on the other (see chapter 5).
Variant Literally, an alternative form. Used in conjunction with locus, phenotype, or mouse strain. A "DNA variant" is equivalent to an alternative DNA allele. A variant mouse usually refers to one that carries a mutant allele or expresses a mutant phenotype.
VNTR "Variable Number of Tandem Repeats" locus; see Minisatellite.
Walking The sequential cloning of adjacent regions along a chromosome by using the ends of previously-obtained clones to re-screen genomic libraries. Walking allows one to extend the length of contigs (see chapter 10).
Wild-type Animal or allele that functions normally and represents a common type found in natural populations at a frequency of at least one percent.
YAC Yeast Artificial Chromosome. A vector for cloning very large genomic inserts of 300 kb to one megabase in length (see chapter 10).
Zygote The fertilized egg containing pronuclei from both the mother and the father.

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