Chapter 8 – Bacterial and Viral Genetic Systems

Bacteria Prototrophic Auxotrophic Wild-type Can grow on minimal media Contains minimal nutrients – carbon, nitrogen, phosphorous, vitamins, ions Auxotrophic Can not produce an essential enzyme or manufacture essential molecules Will only grow on media that contains the “missing” substance Complete media

Culturing bacteria Suspension culture Liquid media Bacteria dies off when nutrients are used up or waste buildup becomes toxic Bacteria grow singularly – no colonies

Culturing bacteria Petri dishes Growth media in agar Isolate individual colonies Each colony originates from a single bacterium

Replica plating Gives “carbon copies” of petri dish colonies Use sterilized velvet to make a stamp Some bacteria from each colony is transferred to velvet, and then transferred to new dishes

Bacterial genome Most consist of a single, circular chromosome Some have several chromosomes, and a few have linear chromosomes Very little “extra” DNA between genes Plasmids Small, circular, extra-chromosomal DNA Usually non-essential Replicate independent of chromosomal DNA Have their own origin of replication

F factor episome Episome A plasmid that can replicate independently AND also has the ability to incorporate into chromosomes

Gene transfer in bacteria - Conjugation One bacteria directly transfers DNA to another bacterium Cytoplasmic connection forms, and either entire plasmid or part of the chromosome is transferred from donor to recipient Crossing over may occur between homolgous regions Creates recombinant DNA Extra DNA is degraded

Gene transfer in bacteria - Transformation Bacteria takes up DNA from surrounding environment Recombination may occur

Gene transfer in bacteria - Transduction Viral particle introduced DNA from a bacterium into a new bacterium

Conjugation Fertility factor/F factor contains ori and genes needed for conjugation F+ and F- F+ contains F factor Forms a sex pilus – extension of cell membrane Extends and comes in contact with F- receptor F factor separates, and one strand is transferred into F- Double stranded DNA is created and F- becomes F+

Conjugation Hfr bacteria F+ cell that has F factor incorporated into chromosome As F factor enters recipient, some chromsome enters – amount depends on time length of contact Donor DNA made into double-stranded Crossing over can occur between homologous regions Any DNA not incorporated is degraded Recipient is not usually converted to F+ since the F factor is nicked in the middle

Conjugation F′ bacteria F factor excises out of a chromosome in a Hfr cell May remove part of chromosome as well F′ plasmid now contains F factor and some genes from chromosome Enters F- bacteria Produces merozygotes – partially diploid

Transformation Uptake of DNA and incorporation into chromosome or plasmid Naturally occurring – dead bacteria Artificially introduced Competent – cells able to take up DNA CaCl2, heat shock, electrical fields Makes membrane more permeable to DNA DNA does not have to have bacterial origin Transformants – bacteria that have incorporated foreign DNA

E. Coli has model organism Many strains are avirulent Small and rapid reproduction Easy to culture Genome is single chromosome - haploid Small genome 4.5 million bp/4,000 genes Wild-type are prototrophic

Viral genetics DNA or RNA (single or double stranded) as genetic material Can not reproduce on their own Bacteriophages – viral particles that infect bacteria

Bacteriophage – lytic cycle Virulent phages Viral DNA is injected into host cell where it replicated, transcribed, and translated into more phages Host cell bursts open to release viral particles

Bacteriophages – lysogenic cycle Temperate phages Phage DNA is incorporated into host genome – prophage Passed onto all progeny cells Can be transcribed and translated Can exit from host genome to enter lytic cycle

Transduction Generalized Any gene is transferred During lytic cycle, bacterial DNA is degraded Some may enter viral protein coat instead of viral genetic material Transducing phages Can become incorporated into new host’s genome

Transduction Specialized Few genes are transferred/genes near certain sites of chromosome During lysogenic cycle, prophage enters at specific sites of host’s genome When prophage excises, it may do so imperfectly and bring some hot DNA with it Then introduced to new host

RNA viruses Positive strand RNA viruses Negative strand RNA viruses Single strand directly codes for viral proteins Negative strand RNA viruses Must make complementary RNA strand, which then codes for proteins Retroviruses Incorporate into host genome Must make DNA from RNA Reverse transcriptase Makes cDNA from DNA or RNA template Enters host genome as a provirus Can be transcribed and translated Some retroviruses contain oncogenes Cause tumors