Ch. 6

All of the following are characteristics of insertion sequences elements

they encode protein they are flanked by inverted repeats there can be more than one copy of an IS element in a bacterial genome

The role of transposase activity in IS element transposition includes

cutting DNA at the target sequence

Which of the following transposition events is most likely to result in a loss of function mutation?

Insertion of an IS element within the coding region of a gene

Virus properties

HIV contains reverse transcriptase. A retrovirus contains RNA. Enveloped viruses bud from the host cell. The capsid enters the host cell if the virus is enveloped.

True or false? A bacterial strain that is pro+ thi+ leu− met− will grow on minimal media plus leucine and thiamine.

False

Which of the following statements about conjugation is true?

DNA is transferred from an F+ cell to an F− cell.

How is a merozygote formed?

The F factor and several adjacent genes are excised from the chromosome of an F+ cell and transferred to an F− strain.

In an interrupted mating experiment, the purpose of plating cells on a selective medium is

to ensure that only recombinant genotypes are recovered

Mapping bacterial genes by conjugation is based on which of the following assumptions?

Genes are transferred from donor to recipient in a linear fashion.

Which of the following statements about mapping bacterial genes by conjugation is NOT true?

The closer a gene to the Hfr origin, the more likely it will be transferred to the recipient during conjugation.

What is the purpose of adding streptomycin to each selection medium?

to ensure that only exconjugate bacteria and not donor bacteria will grow on the media.

Which of the following statements about bacterial genomes is NOT true?

Like eukaryotic chromosomes, bacterial chromosomes are generally linear.

What is the name of the process in bacteria that involves DNA transfer through a cytoplasmic connection known as a pilus?

Conjugation

What is the name of the complex that binds to the origin of transfer (oriT) in the process of conjugation?

Relaxosome

How is an Hfr chromosome formed?

By integration of the F-plasmid into the bacterial chromosome

What map units are used in interrupted mating analyses of conjugation?

minutes

Which of the following conjugation types is a stable partial diploid?

An F’ cell contains an F-plasmid that has picked up a small number of bacterial genes

In which process of genetic exchange in bacteria is one strand of DNA degraded as it enters the recipient cell?

Transformation

What is the name given to a bacteriophage genome integrated into a host cell chromosome?

Prophage

Which process of genetic exchange in bacteria is capable of transferring only a small, specific region of the bacterial chromosome?

Specialized transduction

Conjugation

the transfer of replicated DNA from a donor to a recipient physical union of bacteria, Genetic information is transferred through a hollow tube, called a conjugation pilus or conjugation tube The exconjugant cell is produced by the conjugation

Transformation

uptake of DNA from the environment

Transduction

transfer of DNA from one bacterium to another by a viral vector

Bacterial Genomes

composed of a single chromosome carries mostly essential genes bacterial chromosome = covalently closed circular molecule of ds DNA

Plasmids

small double-stranded circular DNA molecules containing non-essential genes smaller than bacterial chromosomes easily modified

F (fertility) plasmid

contains genes that promote its own transfer from donors to recipients Donor cells possess an F factor and are called F cells whereas recipients, called F cells, lack an F factor

R (resistance) plasmid

carries antibiotic resistance genes that can be transferred to recipient cells

high-copy-number

plasmids that replicate independently q

Low-copy-number

plasmids are present in one or two copies per bacterial cell and usually cannot replicate independently

insertion sequence (IS)

mobile segments of bacterial DNA that can transpose themselves to new locations 4 of them make up F factor

episome

formed when Circular elements like the F factor can integrate into the bacterial chromosome

Mechanism of Conjugation

After contact is established by the conjugation pilus, gene expression from the F factor produces a protein complex called the relaxosome The complex binds the origin of transfer (oriT) on the F factor, and catalyzes the cleavage of one phosphodiester bond on the DNA strand (the T strand) T strand unwinds and a protein called relaxase binds the free 5 end nucleoprotein complex at the 5 end of the T strand provides a recognition signal for the coupling complex nucleoprotein complex binds the coupling complex, then affiliates with the exporter complex that moves the proteins and associated DNA into the recipient cell

rolling circle replication

T strand transfer across the pilus is accompanied by a specialized process called begins at oriT and uses the non-transferred DNA strand as a template for DNA replication

recipient cell

uses the imported DNA as a template for replication

Formation of an Hfr Chromosome

high-frequency recombination F factor integrates into the bacterial chromosome to form the Hfr chromosome occurs rarely

Hfr Gene transfer

Complete transfer of the bacterial chromosome is not accomplished , rolling circle replication Linear DNA in the recipient cell cannot circularize

Selection

Exconjugants are identified by their growth on a selective growth medium

Outcomes of Hfr x F⁻ Mating

Transfer of one or more donor alleles into the recipient chromosome occurs by homologous recombination forms an exconjugant chromosome F factor is not fully transferred during the mating recipient cell is not converted into a donor cell

Interrupted mating

the cessation of conjugation by breaking the conjugation tube

time-of-entry mapping

determine the distances between genes

Limitations of Interrupted Mating

the likelihood of gene transfer drops off quickly Each Hfr strain can transfer genes in one direction only Multiple Hfr strains must be used in interrupted mating experiments to map all of the genes in a species

F factor integration

take place at any IS element on a bacterial chromosome

F’ donor

contains a functional F factor derived from imperfect excision of the F factor from an Hfr chromosome contains all its own DNA plus a segment of the bacterial chromosome

partial diploids

Exconjugants that contain a complete F’ factor they contain two copies of the bacterial chromosome genes found on the F’ factor

Bacterial Transformation

occurs when a recipient cell takes up a fragment of donor DNA from the surrounding growth medium usually shorter than 100 kb useful for mapping genes that are closely linked

cotransformed

transferred into the recipient on the same fragment of DNA

lysis

breakage of a donor cell

Steps in Transformation

1) Donor DNA binds at the receptor site. one strand is degraded as it enters the recipient cell 2) the transforming strand pairs with the homologous region of the recipient chromosome 3) the transforming strand displaces a recipient strand, forming complementary heteroduplex DNA, the excess strand degrades 4) DNA replication and cell division produce one transform ant and one nontransformant

Cotransformation

Closely linked genes will often end up on the same fragment of transforming DNA and require only two crossover events For two genes too far apart to be cotransformed, their cotransformation frequency will be the product of the two transformation frequencies-and will require four crossover events

Bacterial Transduction

transfer of genetic material from a donor to a recipient cell by way of a bacteriophage bacteriophage infects a donor cell and some progeny phages accidentally incorporate a fragment of donor DNA rather than a copy of the phage chromosome

lytic cycle

1. Attachment of the phage to the host cell 2. Injection of the phage chromosome into the host, followed by circularization of the phage chromosome 3. Replication of phage DNA using host proteins and enzymes 4. Transcription and translation of phage genes, and subsequent production of heads, sheaths, and tail fibers for assembly of progeny phage 5. Packaging of phage chromosomes into phage heads 6. Lysis of the host cell, and release of progeny phage particles

lysogenic cycle

1. Attachment and Injection are the same as the lytic cycle 2. Integration of the phage chromosome into the host chromosome via recombination at a specific DNA sequence found in both chromosomes 3. Excision of the prophage 4. Resumption of the lytic cycle, beginning with phage-chromosome replication

transductant strain

transfer of genetic material from a donor bacterium to a recipient via a phage leads to production

Two Types of Transduction

Generalized transduction Specialized transduction

Generalized transduction

the transfer of any bacterial genes, is carried out by bacteriophages that cannot distinguish between phage and bacterial DNA during packing of DNA into new phage particles package a random piece of bacterial DNA into progeny phage heads

Specialized transduction

carried out only by temperate phages and the only genes transferred are those close to the integration site

Cotransduction

closer two genes are on the donor chromosome, the more likely they will be transduced to a recipient together

Cotransduction frequency

depends on the distance between two genes can be used to determine the relative order of three or more genes

Which of the following conjugations will produce the highest frequency of recombinants among the progeny?

Hfr F-

How is an F donor cell generated?

By aberrant excision of the F-plasmid from an Hfr chromosome

Which type of gene transfer in bacteria involves the uptake of exogenous DNA from the environment?

Transformation involves the uptake of DNA ?from the environment by a bacterial cell. Once inside the cell, the transforming DNA must be recombined into the host cell chromosome in order to be maintained

Which of the following mechanisms of gene transfer in bacteria can produce recombinants for only a specific region (a few genes) of the bacterial chromosome?

Specialized transducing phage are produced ?by aberrant excision of a prophage from a lysogenized host cell chromosome. Aberrant excision can only transfer genes adjacent to the site where the phage genome is integrated into the host chromosome (attB in E. coli).