Genomic scanning for expressed sequences in Xp21 identifies the glycerol kinase gene

Article metrics

Abstract

Rapid genomic scanning methods are required to identify expressed sequences and we report an efficient, sensitive and specific approach which relies upon hybridization of an amplified, labeled cDNA library to digested cosmid DNA. We identified expressed sequences within a cosmid in the glycerol kinase (GK) “critical region” of Xp21 that had impressive similarity to prokaryotic GKs. We used this genomic sequence information to clone the human hepatic GK cDNA. Independent confirmation of the identity of this gene was obtained by functional complementation of GK deficient E. coli mutants with a construct containing the complete human X–linked GK coding sequence.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

References

  1. 1

    Monaco, A.P. et al. Isolation of candidate cDNAs for portions of the Duchenne muscular dystrophy gene. Nature 323, 646–650 (1986).

  2. 2

    Bird, A.P. CpG-rich islands and the function of DNA methylation. Nature 321, 209–213 (1986).

  3. 3

    Bird, A.P. CpG islands as gene markers in the vertebrate nucleus. Trends Genet. 3, 342–347 (1987).

  4. 4

    Vollrath, D., Davis, R.W., Connelly, C. & Hieter, P. Physical mapping of large DNA by chromosome fragmentation. Proc. natn. Acad. Sci. U.S.A. 85, 6027–6031 (1988).

  5. 5

    Kurnit, D.M. & Seed, B. Improved genetic selection for screening bacteriophage libraries by homologous recombination in vivo. Proc. natn. Acad. Sci. U.S.A. 87, 3166–3169 (1990).

  6. 6

    Hochgeschwender, U., Sutcliffe, J.G. & Brennan, M.B. Construction and screening of a genomic library specific for mouse chromosome 16. Proc. natn. Acad. Sci. U.S.A. 86, 8482–8486 (1989).

  7. 7

    Liu, P., Legerski, R. & Siciliano, M.J. Isolation of human transcribed sequences from human-rodent somatic cell hybrids. Science 246, 813–815 (1989).

  8. 8

    Corbo, L., Maley, J.A., Nelson, D.L. & Caskey, C.T. Direct cloning of human transcripts with HnRNA from hybrid cell lines. Science 249, 652–655 (1990).

  9. 9

    Duyk, J.M., Kim, S., Myers, R.M. & Cox, D.R. Exon trapping: A genetic screen to identify candidate transcribed sequences in cloned mammalian genomic DNA. Proc. natn. Acad. Sci. U.S.A. 87, 8995–8999 (1990).

  10. 10

    Buckler, A.J. et al. Exon amplification: A strategy to isolate mammalian genes based on RNA splicing. Proc. natn. Acad. Sci. U.S.A. 88, 4005–4009 (1991).

  11. 11

    Elvin, P. et al. Isolation of cDNA clones using yeast artificial chromosome probes. Nucl. Acids Res. 18, 3913–3917 (1990).

  12. 12

    Adams, M. et al. Sequence identification of 2,375 human brain genes. Nature 355, 632–634 (1992).

  13. 13

    Saiki, R.K. et al. Enzymatic amplification of beta-globin genomic sequences and restriction site analysis for diagnosis of sickle cell anemia. Science 230, 1350–1354 (1985).

  14. 14

    Southern, E.M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J. molec. Biol. 98, 503–517 (1975).

  15. 15

    McCabe, E.R.B. Disorders of glycerol metabolism. in The Metabolic Basis of Inherited Disease, (eds Scriver, C.R., Beaudet, A.L, Sly, W.S. & Valle, D.) 945–965 (McGraw-Hill, New York, 1989).

  16. 16

    Walker, A.P. et al. A YAC contig in Xp21 containing the adrenal hypoplasia congenita and glycerol kinase deficiency genes. Hum. molec. Genet. 1, 579–585 (1992).

  17. 17

    Worley, K.C. et al. Yeast artificial chromosome cloning in the glycerol kinase and adrenal hypoplasia congenita region of Xp21. Genomics 16, 407–416 (1993).

  18. 18

    Davies, K.E. et al. Fine mapping of glycerol kinase deficiency and adrenal hypoplasia within Xp21 on the short arm of the human X chromosome. Am. J. med. Genet 29, 557–564 (1988).

  19. 19

    Love, D.R., Bloomfield, J.F., Kenwrick, S.J., Yates, J.R.W. & Davies, K.E. Physical mapping distal to the DMD locus. Genomics 8, 106–112 (1990).

  20. 20

    Altschul, S.F., Gish, W., Miller, W., Myers, E.W. & Lipman, D.J. Basic local alignment search tool. J. molec. Biol. 215, 403–410 (1990).

  21. 21

    Pettigrew, D.W., Ma, D.-P., Conrad, C.A. & Johnson, J.R. Escherichia coli glycerol kinase: Cloning and sequencing of the glpK gene and the primary structure of the enzyme. J. Biol. Chem. 263, 135–139 (1988).

  22. 22

    Holmberg, C., Beijer, L., Rutberg, B. & Rutberg, L. Glycerol catabolism in Bacillus subtilis: Nucleotide sequence of the genes encoding glycerol kinase (glpK) and glycerol-3-phosphate dehydrogenase (iglpD). J. gen. Microbiol. 136, 2367–2375 (1990).

  23. 23

    Guigo, R., Knudsen, S., Drake, N. & Smith, T. Prediction of gene structure. J. molec. Biol. 226, 141–157 (1992).

  24. 24

    Genetics Computer Group (GCG), Sequence Analysis Software Package, Version 7.0 (University Research Park, Madison, Wisconsin, 1991).

  25. 25

    Lupski, J.R. et al. Mutational analysis of the Escherichia coli glpFK region with Tn5 mutagenesis and the polymerase chain reaction. J. Bacteriol. 172, 6129–6134 (1990).

  26. 26

    Sargent, C.A. et al. Cloning of the X-linked glycerol kinase deficiency gene and its identification by sequence comparison to the Bacillus subtilis homologue. Hum. molec. Genet. 2, 97–106 (1993).

  27. 27

    Walker, A.P., Muscatelli, F. & Monaco, A.P. Isolation of the human Xp21 glycerol kinase gene by positional cloning. Hum. molec. Genet. 2, 107–114 (1993).

  28. 28

    Mohri, H. & Masaki, J. Glycerokinase and its possible role in glycerol metabolism of bull spermatozoa. J. Reprod. Fertil. 14, 179–194 (1967).

  29. 29

    Brooks, D.E. The interaction of α-chlorohydrin with glycerol kinase. J. Reprod. Fertil. 56, 593–599 (1979).

  30. 30

    McCarrey, J.R. & Thomas, K. Human testis-specific PGK gene lacks introns and possesses characteristics of a processed gene. Nature 326, 501–505 (1987).

  31. 31

    Dahl, H-H.M., Brown, R.M., Hutchison, W.M., Maragos, C. & Brown, G.K. A testis specific form of the human pyruvate dehydrogenase E1 α subunit is coded for by an intronless gene on chromosome 4. Genomics 8, 225–232 (1990).

  32. 32

    Hurley, J.H. et al. Structure of the regulatory complex of Escherichia coli IIIGlc with glycerol kinase. Science 259, 673–677 (1993).

  33. 33

    Seltzer, W.K. et al. Adrenal dysfunction in glycerol kinase deficiency. Biochem. Med. 33, 189–199 (1985).

  34. 34

    Hensleigh, P.A., Moore, W.V., Wilson, K. & Tulchinsky, D., X-linked adrenal hypoplasia. Obstet. Gynecol. 52, 228–232 (1978).

  35. 35

    Sambrook, J., Fritsch, E.F. & Maniatis, T. Molecular Cloning—A Laboratory Manual (Cold Spring Harbor Laboratory Press, Cold Spring Harbor, 1989).

  36. 36

    Feinberg, A.P. & Vogelstein, B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal. Biochem. 132, 6–13 (1983).

  37. 37

    Church, G.M. & Gilbert, W. Genomic sequencing. Proc. natn. Acad. Sci. U.S.A. 81, 1991–1995 (1984).

  38. 38

    Gribskov, M. & Burgess, R.R. Sigma factors from E. coli, B. subtilis, phage SP01 and phage T4 are homologous proteins. Nucl. Acids Res. 14, 6745–6763 (1986).

Download references

Author information

Rights and permissions

Reprints and Permissions

About this article

Further reading