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30 Cloning Genes of Arabidopsis thaliana by Chromosome Walking

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Arabidopsis Protocols

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 82))

Abstract

Chromosome walking is a versatile technique applicable to cloning virtually any gene of interest identifiable by mutations1. The prmciple of this approach consists of using a molecular probe (for instance, a restriction fragment length polymorphism [RFLP] marker) mapping near the target locus to identify genomic clones, particularly those constructed m high capacity vectors The extremities of the cloned insert are recovered and are used m turn as new probes to identify overlappmg clones extending towards the gene. Repeated rounds (or steps) of such screening would eventually yield an overlapping set of clones (contig) bridgmg the mitral RFLP marker and the gene of interestArabidopsis thaliana is particularly suited for isolating specific genes by this approach because of the small size of the haploid content of its genome and the paucity of repetitive DNA 2

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References

  1. Gibson, S. I. and Somerville, C. (1992) Chromosome walking inArabzdopszs thaliana using yeast artifictal chromosomes, in Methods zn Arabidopsis Research (Koncz, C., Chua, N-H, and Schell, J, eds.), World Scientific Pubhshmg, Smgapore, pp. 119–143

    Google Scholar 

  2. Meyerowitz, E. M. (1994) Structure and organization of theArabzdopszs thallana nuclear genome, inArabldopsls (Meyerowitz, E. M and Somerville, C., eds.), Cold Spring Harbor, Cold Spring Harbor Laboratory Press, NY, pp. 21–36

    Google Scholar 

  3. Gnaudat, J., Hauge, B. M, Valon, C, Smalle, J., Party, F, and Goodman, H. M (1992) Isolation of the Arabrdopsis AB13 gene by positional clonmgPlant Cell 4, 1251–1261.

    Google Scholar 

  4. Retter, R S., Williams, J G. K., Feldmann, K A., Rafalski, J A., Tmgey, S. V., and Scolmk, P A. (1992) Global and local genome mapping in Arabzdopszs thalzana by using recombmant Inbred lines and random amplified polymorphic DNAs. Proc Natl Acad Scr USA 89, 1477–1481

    Google Scholar 

  5. Nam, H.-G., Giraudat, J., den Boer, B., Moonan, F., Loos, W.D. B, Hauge, B. M., and Goodman, H M. (1989) Restriction fragment length polymorphism linkage map of Arabldopsls thallana. Plant Cell 1, 699–705

    CAS  Google Scholar 

  6. Chang, C., Bowman, J. L, DeJohn, A. W, Lander, E. S., and Meyerowitz, E. M(1988) Restriction fragment length polymorphtsm linkage map forArabldopszs thaliana. Proc Nat1 Acad. Sci USA 85, 6856–6860.

    Article  CAS  Google Scholar 

  7. Hauge, B. M, Hanley, S M., Cartmhour, S., Cherry, J. M, Goodman, H. M, Koornneef, M., Stam, P, Chang, C, Kempm, S., Medrano, L., and Meyerowitz, E. M (1993) An integrated genetic/RFLP map of the Arabldopsls thalzana genome. Plant J 3, 745–754.

    Article  CAS  Google Scholar 

  8. Lister, C.,and Dean, C. (1993) Recombinant inbred lines for mapping RFLP and phenotyptc markers m Arabzdopsls thalzana Plant J 4,745–750.

    CAS  Google Scholar 

  9. Lister, C and Dean, C (1995) Latest Lister and Dean RI Map Weeds World2,11–18.

    Google Scholar 

  10. Schmidt, R, West, J., Love, K., Lenehan, Z., Lister, C., Thompson, H., Bouchez, D., and Dean, C. (1995) Physical map and organization ofArubldopsls thafiuna chromosome 4.Sczence270, 48–483.

    Google Scholar 

  11. Vijayraghavan, U., Siddtqt, I., and Meyerowitz, E. (1995) Isolation of an 800 kb contiguous DNA fragment encompassing a 3.5-CM region of chromosome 1 m Arabldopsrs using YAC clonesGenome38,817–823

    CAS  Google Scholar 

  12. Putterill, J., Robson, F, Lee, K., and Coupland, G (1993) Chromosome walking with YAC clones m Arabidopszs. isolation of 1700 kb of contrguous DNA onchromosome 5, includmg a 300 kb region containing the flowering-time gene CO. Mol. Gen Genet. 239, 145–157.

    CAS  Google Scholar 

  13. Bent, A F., Kunkel, B. N., Dahlbeck, D., Brown, K L, Schmidt, R., Giraudat, J, Leung, J., and Staskawicz, B J. (1994) RPS2 of Arabzdopszs thalzana a leucme-rich repeat class of plant disease resistance genes Sczence 265, 1856–1860

    Article  CAS  Google Scholar 

  14. Grill, E. and Somervtlle, C. (1991) Constructton and characterization of a yeast artificial chromosome hbrary of Arubzdopszswhich is suitable for chromosome walkingMol Gen. Genet 226, 484–490.

    Article  PubMed  CAS  Google Scholar 

  15. Ward, E. R and Jen, G. C. (1990) Isolation of smgle-copy-sequence clones from a yeast arttficial chromosome library of randomly-shearedArabzdopszs thalmzaDNA Plant MOE Bzol14,561–568.

    Article  CAS  Google Scholar 

  16. Matallana, E, Bell, C. J., Dunn, P. J, Lu, M, and Ecker, J. R. (1992) Genetic and physical linkage of theArabzdopszs genome: methods for anchormg yeast arttfictal chromosomes, inMethods zn Arabrdopsis Research (Koncz, C., Chua, N-H., and Schell, J.,eds.), World Scienttfic Pubhshmg, Smgapore, pp 144–169.

    Google Scholar 

  17. Creusot, F, Foutlloux, E., Dron, M., Lafeurtel, J, Ptcard, G., Btllault, A., Paslier, D L., Cohen, D., Chaboute, M, Durr, A., Fleck, J., Gigot, C, Camilleri, C., Bellnn, C, Caboche, M., and Bouchez, D (1995) The CIC library: a large Insert DNA ltbrary for genome mapping inArabzdopszsthalzana Plant J 8, 763–770

    CAS  Google Scholar 

  18. Chat, S., Creelman, R A., Mullet, J. E, and Wing, R. A (1995) Construction and characterization of a bacterial artiflctal chromosome library of Arabzdopszsthalzana Plant Mol. Bzol Rep. 13, 124–128.

    Article  Google Scholar 

  19. Liu, Y., Mitsukawa, N., Vazquez-Tello, A.,and Whittier, R F (1995) Generation of a high-quality PI library ofArubidopszssuitable for chromosome walking.Plant J 7,351–358.

    Article  CAS  Google Scholar 

  20. Schmidt, R., Putterill, J, West, J., Cnops, G., Robson, F, Coupland, G, and Dean, C. (1994) Analysts of clones carrying repeated DNA sequences in two YAC libraries ofArabzdopszs thalzanaDNA Plant J. 5,735–744.

    CAS  Google Scholar 

  21. Hwang, I., Kohcht, T., Hauge, B., Goodman, H. M, Schmidt, R., Cnops, G, Dean, C, Gibson, S., Iba, K., Lemteux, B., Arondel, V., Danhoff, L, and Somervtlle, C (1991) Identtfication and map positton of YAC clones comprising one-thud of the Arabidopszs genome. Plant J 1,367–374

    CAS  Google Scholar 

  22. Leung, J., Bouvier-Durand, M, Morris, P.-C, Guerrrer, D., Chefdor, F., and Giraudat, J. (1994) Arabzdopsis ABA response gene ABIl: features of a calciummodulated protein phosphatase. Scrence264, 1448–1452

    Article  CAS  Google Scholar 

  23. Steller, H. and Pirrotta, V. (1985) A transposable P vector that confers selectable G418 resistance toDrosophzla larvae. EMBO J 4, 167–171.

    CAS  Google Scholar 

  24. Sambrook, J., Fntsch, E. F, and Maniatis, T. (1989) Molecular Cloning* A Laboratory Manual, 2d ed. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY

    Google Scholar 

  25. Church, G. M and Gilbert, W (1984) Genomic sequencing. Proc Nut1 Acad.Scz USA 81, 1991–1995.

    Article  CAS  Google Scholar 

  26. Collms, F. S. and Werssman, S. M. (1984) Directional clonmg of DNA fragmentsat alarge distance from an initial probe: a circularization method.Proc Natl Acad Sa L/&4 81, 6812–6816

    Article  Google Scholar 

  27. Boyle, J. S. and Lew, A M. (1995) An inexpensive alternative to glassmilk for DNA purrficatron. Trends Genet. 11,8.

    Article  PubMed  CAS  Google Scholar 

  28. Seed, B., Parker, R. C., and Davrdson, N. (1982) Representation of DNA sequences in recombinant DNA libraries prepared by restriction enzyme partial digestion. Gene 19, 201–209.

    Article  PubMed  CAS  Google Scholar 

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© 1998 Human press Inc, Totowa, NJ

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Leung, J., Giraudat, J. (1998). 30 Cloning Genes of Arabidopsis thaliana by Chromosome Walking. In: Martinez-Zapater, J.M., Salinas, J. (eds) Arabidopsis Protocols. Methods in Molecular Biology™, vol 82. Humana Press. https://doi.org/10.1385/0-89603-391-0:277

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  • DOI: https://doi.org/10.1385/0-89603-391-0:277

  • Publisher Name: Humana Press

  • Print ISBN: 978-0-89603-391-7

  • Online ISBN: 978-1-59259-268-5

  • eBook Packages: Springer Protocols

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