1Chapter 13Microbial Recombination and Plasmids2Eucaryotic recombination• recombination– process in which one or more nucleic acid molecules are rearranged or combined to produce a new nucleotide sequence• in eucaryotes, usually occurs as the result of crossing-over during meiosisFigure 13.13Bacterial Recombination: General Principles• several types of recombination– general recombination• can be reciprocal or nonreciprocal– site-specific recombination– replicative recombination4Reciprocal general recombination• most common type of recombination• a reciprocal exchange between pair of homologous chromosomes• results from DNA strand breakage and reunion, leading to crossing-over5Figure 13.2Reciprocal general recom-bination6Figure 13.27Nonreciprocal general recombination• incorporation of single strand of DNA into chromosome, forming a stretch of heteroduplex DNA• proposed to occur during bacterial transformationFigure 13.38Site-specific recombination• insertion of nonhomologous DNA into a chromosome• often occurs during viral genome integration into host chromosome– enzymes responsible are specific for virus and its host9Replicative recombination• accompanies replication of genetic material• used by genetic elements that move about the genome10Horizontal gene transfer• transfer of genes from one mature, independent organism (donor) to another (recipient)• exogenote– DNA that is transferred to recipient• endogenote– genome of recipient• merozyogote– recipient cell that is temporarily diploid as result of transfer process11Figure 13.4three mechanisms ofgene transfer12Bacterial Plasmids• small, double-stranded, usually circular DNA molecules• are replicons– have their own origin of replication– can exist as single copies or as multiple copies•curing– elimination of plasmid– can be spontaneous or induced by treatments that inhibit plasmid replication but not host cell reproduction13Bacterial plasmids…• episomes– plasmids that can exist either with or without integrating into chromosome• conjugative plasmids– have genes for pili– can transfer copies of themselves to other bacteria during conjugation1415Fertility Factors• conjugative plasmids• e.g., F factor of E. coli• many are also episomesFigure 13.516Figure 13.7mediated byinsertion sequences(IS)F plasmid integration17Resistance Factors• R factors (plasmids)• have genes for resistance to antibiotics• some are conjugative• usually do not integrate into chromosome18Col plasmids• encode colicin – kills E. coli– a type of bacteriocin• protein that destroys other bacteria, usually closely related species• some are conjugative• some carry resistance genes19Other Types of Plasmids• virulence plasmids– carry virulence genes• e.g., genes that confer resistance to host defense mechanisms• e.g., genes that encode toxins• metabolic plasmids– carry genes for metabolic processes• e.g., genes encoding degradative enzymes for pesticides• e.g., genes for nitrogen fixation20Transposable Elements• transposition– the movement of pieces of DNA around the genome• transposable elements (transposons)– segments of DNA that carry genes for transposition• widespread in bacteria, eucaryotes and archaea21Types of transposable elements• insertion sequences (IS elements)– contain only genes encoding enzymes required for transposition• e.g., transposase• composite transposons– carry genes in addition to those needed for transposition– conjugative transposons• carry transfer genes in addition to transposition genes22Figure 13.8IR = inverted repeats2324The transposition event• usually transposon replicated, remaining in original site, while duplicate inserts at another site• insertion generates direct repeats of flanking host DNA25Figure 13.10Tn3 trans-position26Figure 13.1027Figure 13.9Generation of direct repeats28Effects of transposition• mutation in coding region– e.g., deletion of genetic material• arrest of translation or transcription• activation of genes• generation of new plasmids– e.g., resistance plasmids29Figure 13.11R1 plasmidRTF = resistancetransfer factora conjugativeplasmidsources ofresistance genesare transposons30Bacterial Conjugation• transfer of DNA by direct cell to cell contact• discovered 1946 by Lederberg and TatumFigure 13.1231Figure 13.13The U-tube experimentdemonstrated thatdirect cell to cellcontact wasnecessaryafter incubation,bacteria plated onminimal mediano prototrophs32F+x F–Mating•F+= donor– contains F factor•F–= recipient– does not contain F factor• F factor replicated by rolling-circle mechanism and duplicate is transferred• recipients usually become F+• donor remains F+33Figure 13.14aF+x F–mating34Hfr Conjugation• Hfr strain– donor having F factor integrated into its chromosome• both plasmid genes and chromosomal genes are transferred35Figure 13.14bHfr x F–mating36F′ Conjugation•F′ plasmid– formed by incorrect excision from chromosome– contains ≥ 1 genes from chromosome•F′ cell can transfer F′ plasmid to recipientFigure 13.15aintegrated F factorchromosomal gene37Figure 13.15bF′ x F–mating38DNA Transformation• uptake of naked DNA molecule from the environment and incorporation into recipient in a heritable form• competent cell– capable of taking up DNA• may be important route of genetic exchange in nature39Figure 13.16a40Figure 13.16b41DNA bindingproteinnuclease – nicks and degrades onestrandcompetence-specificproteinStreptococcus pneumoniaeFigure 13.1742Artificial transformation• transformation done in laboratory with species that are not normally competent (e.g., E. coli)• variety of techniques used to make cells temporarily competent– e.g., calcium chloride treatment• makes cells more permeable to DNA43Transduction• transfer of bacterial genes by viruses• virulent bacteriophages– reproduce using lytic life cycle• temperate bacteriophages– reproduce using lysogenic life cycle44Phage lambda life cyclesFigure 13.18prophage=integrated form of viral genome45Generalized Transduction• any part of bacterial genome can be transferred• occurs during lytic cycle• during viral assembly, fragments of host DNA mistakenly packaged into phage head– generalized transducing particle46Figure 13.19Generalized transductionabortive transductants- bacteria with nonintegrated transduced DNA47Specialized