Skip to main content
PMC home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1985 Jun;82(11):3731–3735. doi: 10.1073/pnas.82.11.3731

Processing of donor DNA during Haemophilus influenzae transformation: analysis using a model plasmid system.

M L Pifer, H O Smith
PMCID: PMC397861  PMID: 2987941

Abstract

A plasmid system was used to investigate the processing of donor DNA during transformation of competent Haemophilus influenzae. Using biochemical and genetic methods, we have determined that portion of a donor plasmid molecule that, on average, can become integrated into a homologous recipient plasmid during transformation. Our results show that (i) transformation efficiency decreases linearly with donor DNA length over the range of 11 to 3.5 kilobase pairs, (ii) transformation efficiency decreases exponentially with size for donor molecules less than 3.5 kilobase pairs in length, and (iii) 5'-end label, but not 3'-end label, can be specifically incorporated into the resident homologous region. We present a model for donor processing during entry that encompasses and explains these observations.

Full text

PDF
3731

Images in this article

3734Image
on p.3734
3734Image
on p.3734

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Barany F., Kahn M. E., Smith H. O. Directional transport and integration of donor DNA in Haemophilus influenzae transformation. Proc Natl Acad Sci U S A. 1983 Dec;80(23):7274–7278. doi: 10.1073/pnas.80.23.7274. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Beattie K. L., Wakil A. E., Driggers P. H. Action of restriction endonucleases on transforming DNA of Haemophilus influenzae. J Bacteriol. 1982 Oct;152(1):332–337. doi: 10.1128/jb.152.1.332-337.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bertrand K. P., Postle K., Wray L. V., Jr, Reznikoff W. S. Overlapping divergent promoters control expression of Tn10 tetracycline resistance. Gene. 1983 Aug;23(2):149–156. doi: 10.1016/0378-1119(83)90046-x. [DOI] [PubMed] [Google Scholar]
  4. Birnboim H. C., Doly J. A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res. 1979 Nov 24;7(6):1513–1523. doi: 10.1093/nar/7.6.1513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Cato A., Jr, Guild W. R. Transformation and DNA size. I. Activity of fragments of defined size and a fit to a random double cross-over model. J Mol Biol. 1968 Oct 14;37(1):157–178. doi: 10.1016/0022-2836(68)90080-6. [DOI] [PubMed] [Google Scholar]
  6. Danner D. B., Deich R. A., Sisco K. L., Smith H. O. An eleven-base-pair sequence determines the specificity of DNA uptake in Haemophilus transformation. Gene. 1980 Nov;11(3-4):311–318. doi: 10.1016/0378-1119(80)90071-2. [DOI] [PubMed] [Google Scholar]
  7. Danner D. B., Pifer M. L. Plasmid cloning vectors resistant to ampicillin and tetracycline which can replicate in both E. coli and Haemophilus cells. Gene. 1982 Apr;18(1):101–105. doi: 10.1016/0378-1119(82)90062-2. [DOI] [PubMed] [Google Scholar]
  8. Goodgal S. H. DNA uptake in Haemophilus transformation. Annu Rev Genet. 1982;16:169–192. doi: 10.1146/annurev.ge.16.120182.001125. [DOI] [PubMed] [Google Scholar]
  9. Goodgal S. H., Gromkova R. Separation of specific segments of transforming DNA after treatment with endodeoxyribonuclease. Proc Natl Acad Sci U S A. 1973 Feb;70(2):503–506. doi: 10.1073/pnas.70.2.503. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Goodgal S. H., Postel E. H. On the mechanism of integration following transformation with single-stranded DNA of Hemophilus influenzae. J Mol Biol. 1967 Sep 14;28(2):261–273. doi: 10.1016/s0022-2836(67)80008-1. [DOI] [PubMed] [Google Scholar]
  11. Harris-Warrick R. M., Elkana Y., Ehrlich S. D., Lederberg J. Electrophoretic separation of Bacillus subtilis genes. Proc Natl Acad Sci U S A. 1975 Jun;72(6):2207–2211. doi: 10.1073/pnas.72.6.2207. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Herriott R. M., Meyer E. M., Vogt M. Defined nongrowth media for stage II development of competence in Haemophilus influenzae. J Bacteriol. 1970 Feb;101(2):517–524. doi: 10.1128/jb.101.2.517-524.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Hirt B. Selective extraction of polyoma DNA from infected mouse cell cultures. J Mol Biol. 1967 Jun 14;26(2):365–369. doi: 10.1016/0022-2836(67)90307-5. [DOI] [PubMed] [Google Scholar]
  14. Kahn M. E., Barany F., Smith H. O. Transformasomes: specialized membranous structures that protect DNA during Haemophilus transformation. Proc Natl Acad Sci U S A. 1983 Nov;80(22):6927–6931. doi: 10.1073/pnas.80.22.6927. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Kahn M. E., Smith H. O. Transformation in Haemophilus: a problem in membrane biology. J Membr Biol. 1984;81(2):89–103. doi: 10.1007/BF01868974. [DOI] [PubMed] [Google Scholar]
  16. Lataste H., Claverys J. P., Sicard A. M. Relation between the transforming activity of a marker and its proximity to the end of the DNA particle. Mol Gen Genet. 1981;183(1):199–201. doi: 10.1007/BF00270163. [DOI] [PubMed] [Google Scholar]
  17. Lefevre J. C., Claverys J. P., Sicard A. M. Donor deoxyribonucleic acid length and marker effect in pneumococcal transformation. J Bacteriol. 1979 Apr;138(1):80–86. doi: 10.1128/jb.138.1.80-86.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Litt M., Marmur J., Ephrussi-Taylor H., Doty P. THE DEPENDENCE OF PNEUMOCOCCAL TRANSFORMATION ON THE MOLECULAR WEIGHT OF DEOXYRIBOSE NUCLEIC ACID. Proc Natl Acad Sci U S A. 1958 Feb;44(2):144–152. doi: 10.1073/pnas.44.2.144. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Morrison D. A., Guild W. R. Transformation and deoxyribonucleic acid size: extent of degradation on entry varies with size of donor. J Bacteriol. 1972 Dec;112(3):1157–1168. doi: 10.1128/jb.112.3.1157-1168.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Notani N., Goodgal S. H. On the nature of recombinants formed during transformation in Hemophilus influenzae. J Gen Physiol. 1966 Jul;49(6):197–209. doi: 10.1085/jgp.49.6.197. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Repine J. E., Pfenninger O. W., Talmage D. W., Berger E. M., Pettijohn D. E. Dimethyl sulfoxide prevents DNA nicking mediated by ionizing radiation or iron/hydrogen peroxide-generated hydroxyl radical. Proc Natl Acad Sci U S A. 1981 Feb;78(2):1001–1003. doi: 10.1073/pnas.78.2.1001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Smith H. O., Danner D. B., Deich R. A. Genetic transformation. Annu Rev Biochem. 1981;50:41–68. doi: 10.1146/annurev.bi.50.070181.000353. [DOI] [PubMed] [Google Scholar]
  23. Stuy J. H. Fate of transforming DNA in the Haemophilus influenzae transformation system. J Mol Biol. 1965 Sep;13(2):554–570. doi: 10.1016/s0022-2836(65)80117-6. [DOI] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES