Conjugation mapping

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Conjugation Mapping- A part of genetics

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<ul><li> 1. CONJUGATION MAPPING OFE.COLI CHROMOSOMESZULEKHA KHALILMSc BIOCHEMISTRYFERGUSSON COLLEGE, PUNE</li></ul> <p> 2. SOME TERMS5 Types bacterial plasmid:1. F plasmid-Contains tra-gene responsible for conjugation2. R plasmid-Carry genes for resistance to antibiotics3. Col plasmid-Contain genes that code for Colicins; proteinsthat can kill other bacteria4. Degradative plasmid-Enables digestion of unusualsubstance5. Virulence plasmid-Turn the bacterium into a pathogen 3. Contd. Fertility factor: Allows genes to be transferred from onebacterium carrying the F-factor to another bacteriumlacking the F-factor. F factor is a self-replicating, circular DNA plasmid (1/40the size of the main chromosome). F+ Bacteria: Possess F-factor as a plasmid independent ofbacterial genome. F- cell: They do not contain F-factor. They act as recipientof F-factor. Hfr cell: An Hfr cell is a bacterium with a conjugativeplasmid(F-factor) integrated into its genomic DNA. 4. INTRODUCTIONBACTERIAL SEXUAL PROCESSES Eukaryotes have the processes of meiosis to reduce diploidsto haploids, and fertilization to return the cells to thediploid state. Bacterial sexual processes are not so regular. However,they serve the same aim: to mix the genes from twodifferent organisms together. The three bacterial sexual processes: 1. Transformation: Naked DNA is taken up from theenvironment by bacterial cells. 2. Conjugation: Direct transfer of DNA from onebacterial cell to another. 3. Transduction: Use of a bacteriophage (bacterial virus)to transfer DNA between cells. 5. TRANSFORMATION Very important for recombinant DNA work. The essence of recombinant DNA technology is to removeDNA from cells, manipulate it in the test tube, then put itback into living cells. In most cases this is done bytransformation. In the case of E. coli, cells are made competent to betransformed by treatment with calcium ions and heat shock.E. coli cells in this condition readily pick up DNA from theirsurroundings and incorporate it into their genomes. 6. CONJUGATION Meaning: the formation or existence of a link betweenthings. The temporary union of two bacteria or unicellularorganisms for the exchange of genetic material. Was first discovered by Joshua Lederberg &amp; EdwardTatum. William Hayes (1953) demonstrated that genetic exchangein E. coli occurs in only one direction. Genetic transfer is mediated by sex factor F. F is a self-replicating, circular DNA plasmid (1/40 the sizeof the main chromosome). The ability to conjugate is conferred by the F plasmid. 7. Contd.. F+ cells grow special tubes called sex pilli from theirbodies. When an F+ cell bumps into an F- cell, the sexpilli holds them together, and a copy of the F plasmid istransferred from the F+ to the F-. Now both cells areF+. F plasmid contains an origin sequence (O), whichinitiates DNA transfer. It also contains genes for hair-likecell surface (F-pili or sex-pili), which aid in contactbetween cells. 8. MECHANISMFigs. A and BTransfer of the Ffactor 9. Contd.. No conjugation can occur between cells of the same matingtype. Conjugation begins when the F-plasmid is nicked at theorigin, and a single strand is transferred using the rollingcircle mechanism. When transfer is complete, both cells are F + double-stranded. 10. RELAXASE AND RELAXOSOME When conjugation is initiated, via a mating signal, a relaxaseenzyme (an endonuclease) creates a nick in one plasmid DNAstrand at the Origin of transfer, or oriT. The relaxase may work alone or in a complex of over a dozenproteins, known collectively as a relaxosome. The transferred, or T-strand, is unwound from the duplexplasmid and transferred into the recipient bacterium in a5'-terminus to 3'-terminus direction. The remaining strand is replicated, either independent ofconjugative action or in concern with conjugation. 11. MECHANISM ContdEach F+bacterium contains F piliBinding of F pili to specific outer membrane proteinIntercellular cytoplasmic bridge formationTransfer of single strand from donor to recipientTransferred strand isconverted into double strandedcircle in recipient.The copy is retainedin donorThe newly synthesized double strandedDNA is called exogenote and native DNAas endogenote. 12. CONJUGATION OF Hfr CELLS A high-frequency recombination cell (Hfr cell) (also calledan Hfr strain) is a bacterium with a conjugativeplasmid(often the F-factor) integrated into its genomicDNA. No chromosomal DNA is transferred by standard sex factorF. Transfer of chromosome DNA is facilitated by specialstrains of F + integrated into the bacterial chromosome bycrossing over. 13. Contd Hfr strains = high frequency recombination strains. Discovered by William Hayes and Luca Cavalli-Sforza. Hfr strains replicate F -factor as part of their mainchromosome. Conjugation in Hfr strains begins when F + is nicked at theorigin, and F+ and bacteria chromosomal DNA aretransferred using the rolling circle mechanism 14. MECHANISMFig.Transfer of the Hfr F+factor 15. CHARACTERISTICS DNA transfer requires cell-cell contact. DNA transfer occurs via a conjugal pore. DNA transfer occurs in one direction - from donor torecipient not vice versa DNA transfer does not require protein synthesis in donor. DNA transfer requires energy in donor cell - primarily ATP. 16. CONCLUSION The genetic information transferred is often beneficialto the recipient cell. Benefits may include antibioticresistance, other xenobiotic tolerance, or the ability toutilize a new metabolite. Such beneficial plasmids maybe considered bacterial endosymbionts. Some conjugative elements may also be viewed asgenetic parasites on the bacterium, and conjugation asa mechanism was evolved by the mobile element tospread itself into new hosts 17. REFERENCEShttp://en.wikipedia.org/www.ncbi.nlm.nih.govwww.youtube.comAND GOOGLE 18. THANK YOU!! </p>