Abstract
Competent Haemophilus cells recognize and preferentially take up Haemophilus DNA during genetic transformation. This preferential uptake is correlated with the presence on incoming DNA of an 11-base-pair (bp) sequence, 5'-A-A-G-T-G-C-G-G-T-C-A-3'. To prove that this sequence is the recognition site that identifies Haemophilus DNA to the competent cell, we have now constructed a series of plasmids, each of which contains the 11-bp sequence. Using two different assay systems we have tested the ability of fragments from these plasmids to compete with cloned Haemophilus DNA fragments that naturally contain the 11-bp sequence. We find that the addition of the 11-bp sequence to a DNA fragment is necessary and sufficient for preferential uptake of that fragment. However, plasmid DNAs containing this sequence may vary as much as 48-fold in uptake activity, and this variation correlates with the A+T-richness of the DNA flanking the 11-mer.
Full text
PDF![2393](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/71e0/346200/03e94924d142/pnas00446-0263.png)
![2394](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/71e0/346200/748082bde78a/pnas00446-0264.png)
![2395](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/71e0/346200/f2c9c2264e6b/pnas00446-0265.png)
![2396](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/71e0/346200/02ef32efabf5/pnas00446-0266.png)
![2397](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/71e0/346200/77718805016b/pnas00446-0267.png)
Images in this article
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- 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]
- Bochner B. R., Huang H. C., Schieven G. L., Ames B. N. Positive selection for loss of tetracycline resistance. J Bacteriol. 1980 Aug;143(2):926–933. doi: 10.1128/jb.143.2.926-933.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Caster J. H., Goodgal S. H. Competence Mutant of Haemophilus influenzae with Abnormal Ratios of Marker Efficiencies in Transformation. J Bacteriol. 1972 Oct;112(1):492–502. doi: 10.1128/jb.112.1.492-502.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 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]
- Deich R. A., Smith H. O. Mechanism of homospecific DNA uptake in Haemophilus influenzae transformation. Mol Gen Genet. 1980 Feb;177(3):369–374. doi: 10.1007/BF00271475. [DOI] [PubMed] [Google Scholar]
- 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]
- Maxam A. M., Gilbert W. Sequencing end-labeled DNA with base-specific chemical cleavages. Methods Enzymol. 1980;65(1):499–560. doi: 10.1016/s0076-6879(80)65059-9. [DOI] [PubMed] [Google Scholar]
- Narang S. A., Hsiung H. M., Brousseau R. Improved phosphotriester method for the synthesis of gene fragments. Methods Enzymol. 1979;68:90–98. doi: 10.1016/0076-6879(79)68008-4. [DOI] [PubMed] [Google Scholar]
- Nelson T., Brutlag D. Addition of homopolymers to the 3'-ends of duplex DNA with terminal transferase. Methods Enzymol. 1979;68:41–50. doi: 10.1016/0076-6879(79)68005-9. [DOI] [PubMed] [Google Scholar]
- Peck L. J., Wang J. C. Sequence dependence of the helical repeat of DNA in solution. Nature. 1981 Jul 23;292(5821):375–378. doi: 10.1038/292375a0. [DOI] [PubMed] [Google Scholar]
- Rhodes D., Klug A. Sequence-dependent helical periodicity of DNA. Nature. 1981 Jul 23;292(5821):378–380. doi: 10.1038/292378a0. [DOI] [PubMed] [Google Scholar]
- Scocca J. J., Poland R. L., Zoon K. C. Specificity in deoxyribonucleic acid uptake by transformable Haemophilus influenzae. J Bacteriol. 1974 May;118(2):369–373. doi: 10.1128/jb.118.2.369-373.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Seeman N. C., Rosenberg J. M., Rich A. Sequence-specific recognition of double helical nucleic acids by proteins. Proc Natl Acad Sci U S A. 1976 Mar;73(3):804–808. doi: 10.1073/pnas.73.3.804. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Siebenlist U., Simpson R. B., Gilbert W. E. coli RNA polymerase interacts homologously with two different promoters. Cell. 1980 Jun;20(2):269–281. doi: 10.1016/0092-8674(80)90613-3. [DOI] [PubMed] [Google Scholar]
- Sisco K. L., Smith H. O. Sequence-specific DNA uptake in Haemophilus transformation. Proc Natl Acad Sci U S A. 1979 Feb;76(2):972–976. doi: 10.1073/pnas.76.2.972. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 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]
- Wallace R. B., Shaffer J., Murphy R. F., Bonner J., Hirose T., Itakura K. Hybridization of synthetic oligodeoxyribonucleotides to phi chi 174 DNA: the effect of single base pair mismatch. Nucleic Acids Res. 1979 Aug 10;6(11):3543–3557. doi: 10.1093/nar/6.11.3543. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wensink P. C., Finnegan D. J., Donelson J. E., Hogness D. S. A system for mapping DNA sequences in the chromosomes of Drosophila melanogaster. Cell. 1974 Dec;3(4):315–325. doi: 10.1016/0092-8674(74)90045-2. [DOI] [PubMed] [Google Scholar]