Environmental and Molecular Mutagenesis 20:81-83 (1992)
Human HPRT Mutant Database: Software for Data Entry and Retrieval Neal F. Cariello, Teresa R. Craft, Harry Vrieling, Albert A. van Zeeland, Thomas Adams, and Thomas R. Skopek Pathology Department (N.F.C., T.R.S.), Environmental Sciences and Engineering (T.R.S., T.R.C.), University of North Carolina, Alliance Technology Corporation (T.A.) Chapel Hill, North Carolina; Department of Radiation Genetics a n d Chemical Mutagenesis, Leiden University, The Netherlands (H. V., A.A.v.Z.) We have developed a computer database containing information on over 1,000 human hypoxanthine guanine phosphoribosyl transferase (HPRT) mutants. Both published and unpublished data are present. The database itself is maintained in a dBASE format (.DBF) and we provide a set of programs to examine and extract information from the database. A program to input information into the database is also supplied. The dotabase and programs are available directly from us or via remote FTP (file transfer protocol) using BITNETANTERNET. All programs require an IBM-compatible computer, the MS-DOS operating system (version 3.3 or greater), and a hard disk with about 5 megabytes of free disk space.
The purpose of the database is 1) to allow investigators to contribute their HPRT mutants directly to the database in a standardized foshion, and 2) to allow access to the entire database with a set of programs that allows manipulation and extraction of data. For example, using our programs it is possible to i) order the datobose by base pair position, ii)examine only information regarding mutogenesis by a particular agent, iii) search for a particular author, iv) create a report which contains selected portions of the database, the report can be printed or saved as a file. The database will be updated every several months and distributed. Q 1992 WiIey-Liss, Inc.
Key words: hypoxanthine guanine phosphoribosyl transferase, computer database
INTRODUCTION
The hypoxanthine guanine phosphoribosyl transferase gene (HPRT) has been a focus of mutagenesis studies for several decades [for review see Caskey and Kruh, 1979; Fenwick, 19851. Several features of the HPRT gene have made it amenable to study. The gene is X-linked, therefore only a single functional copy exists per cell [Shapiro et al., 19661. In contrast to many autosomal alleles, a single mutational event can produce a mutant cell. The gene is nonessential for cells in culture, and forward and reverse selection systems exist so that both HPRT+ and HPRT- cells can be selected in culture [Szybalski and Szybalski, 1962; Szybalski, 19581. In humans, severe HPRT deficiency can produce the Lesch-Nyhan syndrome [Seegmiller et al., 19671 and a partial HPRT deficiency may lead to severe gout and the overproduction of uric acid [Kelley et al., 19671. It is possible to clone human HPRT- T-cells directly from peripheral blood [Albertini et al., 1985; Turner et al., 19851, thus permitting analysis of mutations arising in vivo in humans. The sequence of the human HPRT cDNA has been available for nearly a decade [Jolly et al., 19831 and recently the entire gene has been sequenced [Edwards et al., 19901.
0 1992 Wiley-Liss, Inc.
Importantly, the polymerase chain reaction has permitted rapid sequence analysis of the HPRT cDNA [Simpson et al., 1988; Vrieling et al., 1988; Yang et al., 19891 and of individual HPRT exons [Cariello et al., 1988; Gibbs et al., 19901. Sequence information about HPRT mutants has been accumulating at a rapid rate in recent years. In an effort to provide tools to catalog and study these mutants, we have developed a HPRT mutant database with software to facilitate data entry and, more importantly, data manipulation and retrieval. RESULTS A N D DISCUSSION
All programs require an IBM-compatible personal computer, the MS-DOS operating system (version 3.3 or higher), and a hard disk with about 5 megabytes of free disk space. Use of a mouse is very strongly recommended for the DATA-OUT program.
Received February 5 , 1992; revised and accepted May 19, 1992. Address reprint requests to Neal F. Cariello, University of North Carolina, Pathology Department, CB# 7525, Chapel Hill, NC 27599.
82
Cariello et al.
Database
The database itself is in a dBASE format (.DBF), so users with access to dBASE, FoxBase+, FoxPro, Clipper, or dBASE-compatible software can examine the database with the program of their choice. However, we also provide programs for data extraction and manipulation, so users are not required to purchase any software in order to use the database. The name of the database file is HUMHPRT.DBF (for HUMan HPRT). The database has over 1,000 entries, and contains both pu6lished and unpublished information. Over 25 different topics of information, called fields, exist for each entry, providing information about the mutagen, dose, nature of the mutagen, sequence information, amino acid alteration, authors, citation, and the like. Logical (true or false) fields are also present, which facilitate the extraction of entire classes of information (for example, all in vivo data). We expect to update the database every several months. DATA-IN Program
DATA-IN.EXE is a program designed to standardize and facilitate data entry into the database. The program prompts the user for information about the nature of the experiment, nature of the mutation, and the like. Help is available at all points in the program by pressing the F1 key. At the end of the program, an ASCII file (normal text, no special characters) which contains the information is written to the hard disk. The experimenter then sends the file to us, preferably by electronic mail, and this information will be added to the database. Information about data submission is provided in the program. DATA-OUT Program
DATA-0UT.EXE is a program that is used to examine information in the database. It is possible to i) search the database for a particular piece of information (e.g., a particular author), ii) order the entire database by information field (e.g., mutagen or base pair position), iii) filter the database so that just the information about a particular topic is visible (e.g., MNNG-induced mutation or in vitro mutations), and iv) create a report with the desired information (the report can be sent to a file or directly to a printer). On-line help is available in the program by pressing the F1 key. The on-line help also contains i) the cDNA sequence for HPRT with numbered base pair positions and numbered amino acid positions, ii) several hundred bases of intron flanking sequence surrounding each exon with numbering according to Edwards et al. [ 19901, iii) information about the contents of each field and iv) important information about getting started. The main method for viewing data is the Browse window. The Browse window is moveable and sizeable, and contains two scroll bars for rapid movement through the database. In
addition, i) the Browse window may be split into two parts with linked or unlinked partitions: (this permits examination of two different parts of the database simultaneously); and ii) Browse fields may be moved and sized (this permits arranging the desired fields side-by-side). Considerable flexibility exists in the creation of reports. Reports are used to obtain a hard copy of information selected from the database. Reports can be printed directly or stored in a file. Fields can be placed where desired on the report page by simply dragging with the mouse. Fields can also be deleted, in this case they do not appear in the report. The user may add lines, boxes, and text to the report. A page preview function also exists so the layout of the report can be examined before printing. Printer driver support for the major printers is provided. Miscellaneous features include a calculator, daily calendar, clock, and puzzle. The user also has the ability to toggle between 25-line mode and 43- or 50-line mode. Experimenters are strongly urged to use this program with a mouse, users without a mouse will be disappointed. The DATA-OUT.EXE and associated files will require about 5 megabytes of hard disk space. Access
The preferred method of obtaining the programs is through BITNET/INTERNET and we have created an account for this purpose. The files are available for transfer using l T P (file transfer protocol), however, the files are binary and must be transferred as such. See your system manager for the details of FTP binary file transfer at your particular site. The files can be accessed via FTP with username SKOPEK and password DATABASE. The node name is UNCVXl and the routing number is 128.109.157.1. A READ.ME file with pertinent information should be read before beginning the file transfer. FTP is a privilege and we ask for your cooperation in maintaining the integrity of our files. The programs will also be available on 720K, 1.2M, and 1.44M floppy disks for users without FTP access. Due to the large size of some of the programs, it is not possible to distribute the programs on 360K floppy disks. Correspondence
The preferred method of communication is EMAIL to CARIELLO@UNCVX 1.BITNET. Postal mail should be addressed to the corresponding author. Please include your name, US postal address, phone, and FAX number in your first communication; this may aid in establishing the proper EMAIL connections. Any comments or suggestions about the database and associated programs will be greatly appreciated. Future Developments
The database itself, HUMHPRT.DBF, will be periodically updated and made available via FTP. Users can then
HPRT Database
simply download the new database and use it with their existing DATA-OUT software. An electronic mailing list will be created to keep users informed of new database and software releases. To be added to the mailing list, send EMAIL stating that you wish to be added to the mailing list. We are in the process of developing software for statistical analysis of the database. This software will be made available when completed.
ACKNOWLEDGMENTS We acknowledge the help of many experimenters in the field, including Richard Albertini, Jane Cole, Irene Jones, Veronica Maher, and Les Recio. The software is made possible by a grant to NFC from the Merck Sharp & Dohme Research Laboratories. NFC is supported in part by NRSA ES-05534-01 and ACS grant PDT423.
REFERENCES Albertini RJ, O’Neill JP, Nicklas JA, Heinz NH, Kelleher PC (1985): Alterations of the hprr gene in human in vivo-derived 6-thioguanine-resistant T lymphocytes. Nature 3 15:369-371. Cariello NF, Scott JK, Kat AG, Thilly WG, Keohavong P (1988): Resolution of a missense mutant in human genomic DNA by denaturing gradient gel electrophoresis and direct sequencing using in vitro DNA amplification: HPRT,,,,,,. Am J Hum Genet 42:726-734. Caskey CT. Kruh GD (1979): The HPRT locus. Cell 16:1-9. Edwards A, Voss H, Rice P, Civitello A, Stegemann J, Schwager C, Zimmermann J, Erne H, Caskey CT, Ansorge W (1990): Automated DNA sequencing of the human HPRT locus. Genomics 6:593-608. Fenwick RG (1985): The HGPRT system. In Gottesman MM (ed): “Molecular Cell Genetics.” New York: Wiley, pp 333-373.
83
Gibbs RA, Nguyen P, Edwards A, Civitello AB, Caskey CT (1990): Multiplex DNA deletion detection and exon sequencing of the hypoxanthine phosphoribosyl transferase gene in Lesch-Nyhan families. Genomics 7:235-244. Jolly DJ, Okayama H, Berg P, Esty AC, Filpula D, Bohlen P, Johnson GG, Shively JE. Hunkapillar T. Friedmann T (1983): Isolation and characterizaton of a full-length expressible cDNA for human hypoxanthine phosphoribosyl transferase. Proc Natl Acad Sci USA 80:477481. Kelley WN, Rosenbloom FM, Seegmiller JE (1967): A specific enzyme defect in gout associated with overproductionof uric acid. Proc Natl Acad Sci USA 57:1735-1739. Seegmiller JE, Rosenbloom FM, Kelly WN ( I 967): Enzyme defect associated with a sex-linked human neurological disorder and excessive purine synthesis. Science 155:1682-1684. Shapiro SL, Sheppard GL, Dreifus FE, Newcombe DS ( I 966): X-linked recessive inheritance of a syndrome of mental retardation with hyperuricemia. Proc Soc Exp Med 122:609-6 1 I . Simpson D, Crosby RM, Skopek TR (1988): A method for specific cloning and sequencing of human HPRT cDNA for mutation analysis. Biochem Biophys Res Comm 151:487-492. Szybalski W (1958): Resistance to 8-azaguanine as a selectable marker for a human cell line. Microbiol Genet Bull 1630-33. Szybalski W, Szybalski EH (1962): Drug sensitivity as a genetic marker for a human cell line. U Mich Med Bull 28:277-293. Turner DR, Morely AA, Haliandros M, Kutlaca R, Sanderson BJ (1985): In vivo somatic mutations in human lymphocytes frequently result from major gene alterations. Nature 3 I5:343-345. Vrieling H, Simons JWIM, van Zeeland AA (1988): Nucleotide sequence determination of point mutations at the mouse HPRT locus using in vitro amplification of HPRT mRNA sequences. Mutat Res 198:107-113. Yang JL, Maher VM, McCormick JJ (1989): Amplification and direct sequencing of cDNA from the lysate of low numbers of diploid human cells. Gene 83:347-354.
Accepted byJ.P. O’Neill
Human HPRT Mutant Database: Software for Data Entry and Retrieval Neal F. Cariello, Teresa R. Craft, Harry Vrieling, Albert A. van Zeeland, Thomas Adams, and Thomas R. Skopek Pathology Department (N.F.C., T.R.S.), Environmental Sciences and Engineering (T.R.S., T.R.C.), University of North Carolina, Alliance Technology Corporation (T.A.) Chapel Hill, North Carolina; Department of Radiation Genetics a n d Chemical Mutagenesis, Leiden University, The Netherlands (H. V., A.A.v.Z.) We have developed a computer database containing information on over 1,000 human hypoxanthine guanine phosphoribosyl transferase (HPRT) mutants. Both published and unpublished data are present. The database itself is maintained in a dBASE format (.DBF) and we provide a set of programs to examine and extract information from the database. A program to input information into the database is also supplied. The dotabase and programs are available directly from us or via remote FTP (file transfer protocol) using BITNETANTERNET. All programs require an IBM-compatible computer, the MS-DOS operating system (version 3.3 or greater), and a hard disk with about 5 megabytes of free disk space.
The purpose of the database is 1) to allow investigators to contribute their HPRT mutants directly to the database in a standardized foshion, and 2) to allow access to the entire database with a set of programs that allows manipulation and extraction of data. For example, using our programs it is possible to i) order the datobose by base pair position, ii)examine only information regarding mutogenesis by a particular agent, iii) search for a particular author, iv) create a report which contains selected portions of the database, the report can be printed or saved as a file. The database will be updated every several months and distributed. Q 1992 WiIey-Liss, Inc.
Key words: hypoxanthine guanine phosphoribosyl transferase, computer database
INTRODUCTION
The hypoxanthine guanine phosphoribosyl transferase gene (HPRT) has been a focus of mutagenesis studies for several decades [for review see Caskey and Kruh, 1979; Fenwick, 19851. Several features of the HPRT gene have made it amenable to study. The gene is X-linked, therefore only a single functional copy exists per cell [Shapiro et al., 19661. In contrast to many autosomal alleles, a single mutational event can produce a mutant cell. The gene is nonessential for cells in culture, and forward and reverse selection systems exist so that both HPRT+ and HPRT- cells can be selected in culture [Szybalski and Szybalski, 1962; Szybalski, 19581. In humans, severe HPRT deficiency can produce the Lesch-Nyhan syndrome [Seegmiller et al., 19671 and a partial HPRT deficiency may lead to severe gout and the overproduction of uric acid [Kelley et al., 19671. It is possible to clone human HPRT- T-cells directly from peripheral blood [Albertini et al., 1985; Turner et al., 19851, thus permitting analysis of mutations arising in vivo in humans. The sequence of the human HPRT cDNA has been available for nearly a decade [Jolly et al., 19831 and recently the entire gene has been sequenced [Edwards et al., 19901.
0 1992 Wiley-Liss, Inc.
Importantly, the polymerase chain reaction has permitted rapid sequence analysis of the HPRT cDNA [Simpson et al., 1988; Vrieling et al., 1988; Yang et al., 19891 and of individual HPRT exons [Cariello et al., 1988; Gibbs et al., 19901. Sequence information about HPRT mutants has been accumulating at a rapid rate in recent years. In an effort to provide tools to catalog and study these mutants, we have developed a HPRT mutant database with software to facilitate data entry and, more importantly, data manipulation and retrieval. RESULTS A N D DISCUSSION
All programs require an IBM-compatible personal computer, the MS-DOS operating system (version 3.3 or higher), and a hard disk with about 5 megabytes of free disk space. Use of a mouse is very strongly recommended for the DATA-OUT program.
Received February 5 , 1992; revised and accepted May 19, 1992. Address reprint requests to Neal F. Cariello, University of North Carolina, Pathology Department, CB# 7525, Chapel Hill, NC 27599.
82
Cariello et al.
Database
The database itself is in a dBASE format (.DBF), so users with access to dBASE, FoxBase+, FoxPro, Clipper, or dBASE-compatible software can examine the database with the program of their choice. However, we also provide programs for data extraction and manipulation, so users are not required to purchase any software in order to use the database. The name of the database file is HUMHPRT.DBF (for HUMan HPRT). The database has over 1,000 entries, and contains both pu6lished and unpublished information. Over 25 different topics of information, called fields, exist for each entry, providing information about the mutagen, dose, nature of the mutagen, sequence information, amino acid alteration, authors, citation, and the like. Logical (true or false) fields are also present, which facilitate the extraction of entire classes of information (for example, all in vivo data). We expect to update the database every several months. DATA-IN Program
DATA-IN.EXE is a program designed to standardize and facilitate data entry into the database. The program prompts the user for information about the nature of the experiment, nature of the mutation, and the like. Help is available at all points in the program by pressing the F1 key. At the end of the program, an ASCII file (normal text, no special characters) which contains the information is written to the hard disk. The experimenter then sends the file to us, preferably by electronic mail, and this information will be added to the database. Information about data submission is provided in the program. DATA-OUT Program
DATA-0UT.EXE is a program that is used to examine information in the database. It is possible to i) search the database for a particular piece of information (e.g., a particular author), ii) order the entire database by information field (e.g., mutagen or base pair position), iii) filter the database so that just the information about a particular topic is visible (e.g., MNNG-induced mutation or in vitro mutations), and iv) create a report with the desired information (the report can be sent to a file or directly to a printer). On-line help is available in the program by pressing the F1 key. The on-line help also contains i) the cDNA sequence for HPRT with numbered base pair positions and numbered amino acid positions, ii) several hundred bases of intron flanking sequence surrounding each exon with numbering according to Edwards et al. [ 19901, iii) information about the contents of each field and iv) important information about getting started. The main method for viewing data is the Browse window. The Browse window is moveable and sizeable, and contains two scroll bars for rapid movement through the database. In
addition, i) the Browse window may be split into two parts with linked or unlinked partitions: (this permits examination of two different parts of the database simultaneously); and ii) Browse fields may be moved and sized (this permits arranging the desired fields side-by-side). Considerable flexibility exists in the creation of reports. Reports are used to obtain a hard copy of information selected from the database. Reports can be printed directly or stored in a file. Fields can be placed where desired on the report page by simply dragging with the mouse. Fields can also be deleted, in this case they do not appear in the report. The user may add lines, boxes, and text to the report. A page preview function also exists so the layout of the report can be examined before printing. Printer driver support for the major printers is provided. Miscellaneous features include a calculator, daily calendar, clock, and puzzle. The user also has the ability to toggle between 25-line mode and 43- or 50-line mode. Experimenters are strongly urged to use this program with a mouse, users without a mouse will be disappointed. The DATA-OUT.EXE and associated files will require about 5 megabytes of hard disk space. Access
The preferred method of obtaining the programs is through BITNET/INTERNET and we have created an account for this purpose. The files are available for transfer using l T P (file transfer protocol), however, the files are binary and must be transferred as such. See your system manager for the details of FTP binary file transfer at your particular site. The files can be accessed via FTP with username SKOPEK and password DATABASE. The node name is UNCVXl and the routing number is 128.109.157.1. A READ.ME file with pertinent information should be read before beginning the file transfer. FTP is a privilege and we ask for your cooperation in maintaining the integrity of our files. The programs will also be available on 720K, 1.2M, and 1.44M floppy disks for users without FTP access. Due to the large size of some of the programs, it is not possible to distribute the programs on 360K floppy disks. Correspondence
The preferred method of communication is EMAIL to CARIELLO@UNCVX 1.BITNET. Postal mail should be addressed to the corresponding author. Please include your name, US postal address, phone, and FAX number in your first communication; this may aid in establishing the proper EMAIL connections. Any comments or suggestions about the database and associated programs will be greatly appreciated. Future Developments
The database itself, HUMHPRT.DBF, will be periodically updated and made available via FTP. Users can then
HPRT Database
simply download the new database and use it with their existing DATA-OUT software. An electronic mailing list will be created to keep users informed of new database and software releases. To be added to the mailing list, send EMAIL stating that you wish to be added to the mailing list. We are in the process of developing software for statistical analysis of the database. This software will be made available when completed.
ACKNOWLEDGMENTS We acknowledge the help of many experimenters in the field, including Richard Albertini, Jane Cole, Irene Jones, Veronica Maher, and Les Recio. The software is made possible by a grant to NFC from the Merck Sharp & Dohme Research Laboratories. NFC is supported in part by NRSA ES-05534-01 and ACS grant PDT423.
REFERENCES Albertini RJ, O’Neill JP, Nicklas JA, Heinz NH, Kelleher PC (1985): Alterations of the hprr gene in human in vivo-derived 6-thioguanine-resistant T lymphocytes. Nature 3 15:369-371. Cariello NF, Scott JK, Kat AG, Thilly WG, Keohavong P (1988): Resolution of a missense mutant in human genomic DNA by denaturing gradient gel electrophoresis and direct sequencing using in vitro DNA amplification: HPRT,,,,,,. Am J Hum Genet 42:726-734. Caskey CT. Kruh GD (1979): The HPRT locus. Cell 16:1-9. Edwards A, Voss H, Rice P, Civitello A, Stegemann J, Schwager C, Zimmermann J, Erne H, Caskey CT, Ansorge W (1990): Automated DNA sequencing of the human HPRT locus. Genomics 6:593-608. Fenwick RG (1985): The HGPRT system. In Gottesman MM (ed): “Molecular Cell Genetics.” New York: Wiley, pp 333-373.
83
Gibbs RA, Nguyen P, Edwards A, Civitello AB, Caskey CT (1990): Multiplex DNA deletion detection and exon sequencing of the hypoxanthine phosphoribosyl transferase gene in Lesch-Nyhan families. Genomics 7:235-244. Jolly DJ, Okayama H, Berg P, Esty AC, Filpula D, Bohlen P, Johnson GG, Shively JE. Hunkapillar T. Friedmann T (1983): Isolation and characterizaton of a full-length expressible cDNA for human hypoxanthine phosphoribosyl transferase. Proc Natl Acad Sci USA 80:477481. Kelley WN, Rosenbloom FM, Seegmiller JE (1967): A specific enzyme defect in gout associated with overproductionof uric acid. Proc Natl Acad Sci USA 57:1735-1739. Seegmiller JE, Rosenbloom FM, Kelly WN ( I 967): Enzyme defect associated with a sex-linked human neurological disorder and excessive purine synthesis. Science 155:1682-1684. Shapiro SL, Sheppard GL, Dreifus FE, Newcombe DS ( I 966): X-linked recessive inheritance of a syndrome of mental retardation with hyperuricemia. Proc Soc Exp Med 122:609-6 1 I . Simpson D, Crosby RM, Skopek TR (1988): A method for specific cloning and sequencing of human HPRT cDNA for mutation analysis. Biochem Biophys Res Comm 151:487-492. Szybalski W (1958): Resistance to 8-azaguanine as a selectable marker for a human cell line. Microbiol Genet Bull 1630-33. Szybalski W, Szybalski EH (1962): Drug sensitivity as a genetic marker for a human cell line. U Mich Med Bull 28:277-293. Turner DR, Morely AA, Haliandros M, Kutlaca R, Sanderson BJ (1985): In vivo somatic mutations in human lymphocytes frequently result from major gene alterations. Nature 3 I5:343-345. Vrieling H, Simons JWIM, van Zeeland AA (1988): Nucleotide sequence determination of point mutations at the mouse HPRT locus using in vitro amplification of HPRT mRNA sequences. Mutat Res 198:107-113. Yang JL, Maher VM, McCormick JJ (1989): Amplification and direct sequencing of cDNA from the lysate of low numbers of diploid human cells. Gene 83:347-354.
Accepted byJ.P. O’Neill
