Biology:DNA–DNA hybridization

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Short description: Technique used to measure similarity in DNA sequences
This page is about the specific use in genomics. For the general phenomenon, see Nucleic acid thermodynamics § Hybridization.

In genomics, DNA–DNA hybridization is a molecular biology technique that measures the degree of genetic similarity between DNA sequences. It is used to determine the genetic distance between two organisms and has been used extensively in phylogeny and taxonomy.[1]

Method

The DNA of one organism is labelled, then mixed with the unlabelled DNA to be compared against. The mixture is incubated to allow DNA strands to dissociate and then cooled to form renewed hybrid double-stranded DNA. Hybridized sequences with a high degree of similarity will bind more firmly, and require more energy to separate them: i.e. they separate when heated at a higher temperature than dissimilar sequences, a process known as "DNA melting".[2][3][4]

To assess the melting profile of the hybridized DNA, the double-stranded DNA is bound to a column or filter and the mixture is heated in small steps. At each step, the column or filter is washed; sequences that melt become single-stranded and wash off. The temperatures at which labelled DNA comes off reflects the amount of similarity between sequences (and the self-hybridization sample serves as a control). These results are combined to determine the degree of genetic similarity between organisms.[5]

One method was introduced for hybridizing large numbers of DNA samples against large numbers of DNA probes on a single membrane. These samples would have to be separated in their own lanes inside the membranes and then the membrane would have to be rotated to a different angle where it would result in simultaneous hybridization with many different DNA probes.[6]

Uses

When several species are compared, similarity values allow organisms to be arranged in a phylogenetic tree; it is therefore one possible approach to carrying out molecular systematics.[citation needed]

In microbiology

DNA–DNA hybridization (DDH) is used as a primary method to distinguish bacterial species as it is difficult to visually classify them accurately.[7] This technique is not widely used on larger organisms where differences in species are easier to identify. In the late 1900s, strains were considered to belong to the same species if they had a DNA–DNA similarity value greater than 70% and their melting temperatures were within 5 °C of each other.[8][9][10] In 2014, a threshold of 79% similarity has been suggested to separate bacterial subspecies.[11]

DDH is a common technique for bacteria, but it is labor intensive, error-prone, and technically challenging. In 2004, a new DDH technique was described. This technique utilized microplates and colorimetrically labelled DNA to decrease the time needed and increase the amount of samples that can be processed.[12] This new DDH technique became the standard for bacterial taxonomy.[13]

In zoology

Charles Sibley and Jon Ahlquist, pioneers of the technique, used DNA–DNA hybridization to examine the phylogenetic relationships of avians (the Sibley–Ahlquist taxonomy) and primates.[14][15]

In radioactivity

In 1969, one such method was performed by Mary Lou Pardue and Joseph G. Gall at the Yale University through radioactivity where it involved the hybridization of a radioactive test DNA in solution to the stationary DNA of a cytological preparation, which is identified as autoradiography.[16]

Replacement by genome sequencing

Critics argue that the technique is inaccurate for comparison of closely related species, as any attempt to measure differences between orthologous sequences between organisms is overwhelmed by the hybridization of paralogous sequences within an organism's genome.[17][better source needed][better source needed] DNA sequencing and computational comparisons of sequences is now generally the method for determining genetic distance, although the technique is still used in microbiology to help identify bacteria.[18]

In silico methods

The modern approach is to carry out DNA–DNA hybridization in silico utilizes completely or partially sequenced genomes.[19] The GGDC and TYGS developed at DSMZ are the most accurate known tools for calculating DDH-analogous values.[19] Among other algorithmic improvements, it solves the problem with paralogous sequences by carefully filtering them from the matches between the two genome sequences. The method has been used for resolving difficult taxa such as Escherichia coli, Bacillus cereus group, and Aeromonas.[20] The Judicial Commission of International Committee on Systematics of Prokaryotes has admitted dDDH as taxonomic evidence.[21]

See also

References

  1. ↑ Erko Stackebrandt (8 September 2010). Molecular Identification, Systematics, and Population Structure of Prokaryotes. Springer Science & Business Media. ISBN 978-3-540-31292-5. https://books.google.com/books?id=eIf6RQeOZPoC. 
  2. ↑ Sinden, Richard R. (1994). DNA structure and function. San Diego: Academic Press. pp. 37–45. ISBN 0-12-645750-6. OCLC 30109829. https://www.worldcat.org/oclc/30109829. 
  3. ↑ Tools and techniques in biomolecular science. Aysha Divan, Janice Royds. Oxford: Oxford University Press. 2013. ISBN 978-0-19-969556-0. OCLC 818450218. https://www.worldcat.org/oclc/818450218. 
  4. ↑ Forster, A. C.; McInnes, J. L.; Skingle, D. C.; Symons, R. H. (1985-02-11). "Non-radioactive hybridization probes prepared by the chemical labelling of DNA and RNA with a novel reagent, photobiotin". Nucleic Acids Research 13 (3): 745–761. doi:10.1093/nar/13.3.745. ISSN 0305-1048. PMID 2582358. 
  5. ↑ Hood, D. W.; Dow, C. S.; Green, P. N. (1987). "DNA:DNA hybridization studies on the pink-pigmented facultative methylotrophs". Journal of General Microbiology 133 (3): 709–720. doi:10.1099/00221287-133-3-709. ISSN 0022-1287. PMID 3655730. https://pubmed.ncbi.nlm.nih.gov/3655730. 
  6. ↑ Socransky, S. S.; Smith, C.; Martin, L.; Paster, B. J.; Dewhirst, F. E.; Levin, A. E. (October 1994). ""Checkerboard" DNA-DNA hybridization". BioTechniques 17 (4): 788–792. ISSN 0736-6205. PMID 7833043. 
  7. ↑ Auch, Alexander F.; von Jan, Mathias; Klenk, Hans-Peter; Göker, Markus (2010). "Digital DNA-DNA hybridization for microbial species delineation by means of genome-to-genome sequence comparison" (in en). Standards in Genomic Sciences 2 (1): 117–134. doi:10.4056/sigs.531120. ISSN 1944-3277. PMID 21304684. 
  8. ↑ Brenner DJ (1973). "Deoxyribonucleic acid reassociation in the taxonomy of enteric bacteria". International Journal of Systematic Bacteriology 23 (4): 298–307. doi:10.1099/00207713-23-4-298. 
  9. ↑ "Report of the ad hoc committee on reconciliation of approaches to bacterial systematics". International Journal of Systematic Bacteriology 37 (4): 463–464. 1987. doi:10.1099/00207713-37-4-463. 
  10. ↑ "Notes on the characterization of prokaryote strains for taxonomic purposes". International Journal of Systematic and Evolutionary Microbiology 60 (Pt 1): 249–266. 2010. doi:10.1099/ijs.0.016949-0. PMID 19700448. 
  11. ↑ "Complete genome sequence of DSM 30083T, the type strain (U5/41T) of Escherichia coli, and a proposal for delineating subspecies in microbial taxonomy". Standards in Genomic Sciences 9: 2. 2013. doi:10.1186/1944-3277-9-2. PMID 25780495. 
  12. ↑ Mehlen, André; Goeldner, Marcia; Ried, Sabine; Stindl, Sibylle; Ludwig, Wolfgang; Schleifer, Karl-Heinz (November 2004). "Development of a fast DNA-DNA hybridization method based on melting profiles in microplates". Systematic and Applied Microbiology 27 (6): 689–695. doi:10.1078/0723202042369875. ISSN 0723-2020. PMID 15612626. 
  13. ↑ Huang, Chien-Hsun; Li, Shiao-Wen; Huang, Lina; Watanabe, Koichi (2018). "Identification and Classification for the Lactobacillus casei Group". Frontiers in Microbiology 9: 1974. doi:10.3389/fmicb.2018.01974. ISSN 1664-302X. PMID 30186277. 
  14. ↑ Genetic Similarities: Wilson, Sarich, Sibley, and Ahlquist
  15. ↑ C.G. Sibley; J.E. Ahlquist (1984). "The Phylogeny of the Hominoid Primates, as Indicated by DNA–DNA Hybridization". Journal of Molecular Evolution 20 (1): 2–15. doi:10.1007/BF02101980. PMID 6429338. Bibcode: 1984JMolE..20....2S. 
  16. ↑ Pardue, Mary Lou, and Joseph G Hall. “Molecular Hybridization of Radioactive DNA to the DNA of Cytological Preparations.” Kline Biology Tower, Yale University, 13 Aug. 1969.
  17. ↑ Marks, Jonathan (2007-05-09). "DNA hybridization in the apes—Technical issues". http://personal.uncc.edu/jmarks/DNAHYB/Dnahyb2.html. 
  18. ↑ S.S. Socransky; A.D. Haffajee; C. Smith; L. Martin; J.A. Haffajee; N.G. Uzel; J. M. Goodson (2004). "Use of checkerboard DNA–DNA hybridization to study complex microbial ecosystems". Oral Microbiology and Immunology 19 (6): 352–362. doi:10.1111/j.1399-302x.2004.00168.x. PMID 15491460. 
  19. ↑ 19.0 19.1 "Genome sequence-based species delimitation with confidence intervals and improved distance functions". BMC Bioinformatics 14: 60. 2013. doi:10.1186/1471-2105-14-60. PMID 23432962. 
  20. ↑ Riojas, Marco A.; McGough, Katya J.; Rider-Riojas, Cristin J.; Rastogi, Nalin; Hazbón, Manzour Hernando (1 January 2018). "Phylogenomic analysis of the species of the Mycobacterium tuberculosis complex demonstrates that Mycobacterium africanum, Mycobacterium bovis, Mycobacterium caprae, Mycobacterium microti and Mycobacterium pinnipedii are later heterotypic synonyms of Mycobacterium tuberculosis". International Journal of Systematic and Evolutionary Microbiology 68 (1): 324–332. doi:10.1099/ijsem.0.002507. PMID 29205127. 
  21. ↑ Arahal, David R.; Bull, Carolee T.; Busse, Hans-Jürgen; Christensen, Henrik; Chuvochina, Maria; Dedysh, Svetlana N.; Fournier, Pierre-Edouard; Konstantinidis, Konstantinos T. et al. (27 April 2023). "Judicial Opinions 123–127". International Journal of Systematic and Evolutionary Microbiology 72 (12). doi:10.1099/ijsem.0.005708. 

Further reading

  • Graur, D. & Li, W-H. 1991 (2nd ed. 1999). Fundamentals of Molecular Evolution.



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