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Microbial Genetics (Micr340)

Microbial Genetics (Micr340) . Lecture 7 Plasmids. Plasmids. DNA molecules other than chromosomes Widely present in most bacteria Roles: adaptation, evolution, pathogenesis Usually encode genes not essential Mostly circular Sizes: a few kb to >100 kb. Naming Plasmids.

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Microbial Genetics (Micr340)

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  1. Microbial Genetics (Micr340) Lecture 7 Plasmids

  2. Plasmids • DNA molecules other than chromosomes • Widely present in most bacteria • Roles: adaptation, evolution, pathogenesis • Usually encode genes not essential • Mostly circular • Sizes: a few kb to >100 kb

  3. Naming Plasmids • Many name after genes they carry or functions of the plasmids • In the beginning: • ColE1, producing Bacteriocin to kill E. coli • Tol, degradation of toluene • Ti, Tumor initiation in plants • RK2, resistance to amp, tet, kanamycin • Now: pXXNNNN. eg, pHX200

  4. Plasmidstructure

  5. Plasmid purification

  6. Plasmid purification

  7. Plasmid replication • Replicons: DNA molecules that can replicate autonomously in cell (chromosome, plasmid) • All replicons have at least one origin of replication (ori) • For plasmid ori, it is called oriV (vegetative) • Two mechanisms of replication: • Theta replication • Rolling-circle replication

  8. Theta replication

  9. Rolling-circle replication

  10. Function of ori region • Replication related genes close to ori. • Host range determination • Narrow host range • Broad host range • Regulation of copy-number • Relaxed plasmids – high copy-number • Stringent plasmids – low copy-number

  11. Regulation of replication: ColE1 plasmids

  12. Random plasmid curing

  13. Plasmid Partition • Plasmid avoid being lost from dividing cells by carrying partitioning systems • The functions involved in these systems are called par functions • cis-acting site: parS • trans-acting site: parA and parB

  14. Incompatibility • Many bacteria contain multiple types of plasmids and can coexist for many generations • Not all types of plasmids can stably coexist in same cell; some interfere with each other’s replication or partition and one of them will be lost • This is called plasmid incompatibility

  15. Incompatibility • Two plasmids that cannot stably coexist are members of the same incompatibility (Inc) group • If two plasmids can stably coexist, they belong to different Inc group

  16. Coexistence of different Inc groups

  17. Coexistence of same Inc groups

  18. Measuring curing of a plasmid

  19. Finding the ori of a plasmid

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