Bacterial gene mapping

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1. Name: Aamer al zaman Roll no: 08 Class : BS Botany 5 th semester Topic: Bacterial gene mapping.

2. What is gene mapping: Genome mapping is the process of describing a genome in terms of the relative locations of genes and other DNA sequences

3. Bacterial gene mapping : Once inside the F −  cell, the linear single-stranded DNA molecule acts as a polymerization template and is converted into a DNA double helix. This linear donor fragment is the exogenote, and the resident F −  chromosome is the endogenote. ... Gene  transfer and recombination provide the key to  mapping  the  bacterial chromosome .

4. Discription: Gene mapping   describes the methods used to identify the  locus  of a gene and the distances between genes . The essence of all  genome  mapping is to place a collection of molecular markers onto their respective positions on the genome.  Molecular markers  come in all forms. Genes can be viewed as one special type of genetic markers in the construction of genome maps, and mapped the same way as any other markers.

5. Genetics mapping via physical mapping: There are two distinctive types of "Maps " used in the field of genome mapping: genetic maps and physical maps. While both maps are a collection of  genetic markers  and gene loci, genetic maps' distances are based on the genetic linkage information, while physical maps use actual physical distances usually measured in number of base pairs

6. Continu: While the physical map could be a more "accurate" representation of the genome, genetic maps often offer insights into the nature of different regions of the chromosome , e.g. the genetic distance to physical distance ratio varies greatly at different genomic regions which reflects different recombination rates, and such rate is often indicative of euchromatic (usually gene-rich) vs heterochromatic (usually gene poor) regions of the genome.

7. Bacterial conjugation: Conjugation is merely the fusion of two compatible bacterial cells. Bringing two genotypes together and allowing them to conjugate is the equivalent of making a  cross  in eukaryotes. Our discussion of  conjugation  will center on the gut bacterium  Escherichia coli (E. coli).  Conjugation and  gene  transfer in E . coli  are driven by a circular  DNA   plasmid  called the  fertility factor  or  sex factor (F),  which is found in some but not all cells. Hence to understand how to make a cross in  E. coli,  we have to understand the properties of F .

8. Recombination between Donor and Recipient DNA: Conjugation allows genes from two different parental cells to come together in the same cell and hence provides an opportunity for  recombination  to occur. Hence mapping analysis is possible. All conjugations (“crosses”) are by definition of the type Hfr (donor) × F −  (recipient). After cell union, the Hfr  chromosome  replicates in a peculiar manner that reels out a single-stranded  DNA  molecule, which is then transferred linearly into the  F −  cell . The  replication  and transfer begin at a specific point at one side of the integrated F, called the  origin (O).  Genes close to the origin are transferred first.

9. Continu: Once inside the  F −  cell , the linear single-stranded  DNA  molecule acts as a polymerization  template  and is converted into a DNA  double helix . This linear donor fragment is the  exogenote , and the resident F − chromosome  is the  endogenote . As a free molecule, the exogenote cannot replicate and will become lost, but because exogenotes and endogenotes are homologous,  crossing-over  can take place between them.  A single crossover between a linear  molecule (the exogenote) and a circular one (the endogenote) would produce a single long molecule that would be inviable. However,  two  crossovers would integrate a part of the donor  genome  into the recipient.

10. Mapping by Interrupted Conjugation: In mapping by interrupted  conjugation , the Hfr and F −  cells are mixed, and conjugation proceeds. Then, at fixed times, the F −  cells are sampled to determine which donor alleles have entered. This sampling is accomplished by using a kitchen blender to separate the joined cells, resulting in interrupted conjugation. After separation, the Hfr cells are selectively killed, and the remaining F −  cells, the  exconjugants,  are tested to see which of the donor alleles have entered and stably recombined with the  endogenote .

11. Continu:  The times at which various donor alleles  first  appear in the exconjugants are calculated. If a donor  allele   a + enters the recipient at 5 minutes after union and allele  b +  enters at 8 minutes, then the two genes are said to be 3 minutes apart on the  chromosome . The map units in this case are minutes. Like the maps based on crossover frequencies, these  linkage  maps are purely genetic constructions. Although the amount of  DNA corresponding to a minute is now known

12. Continu:   A typical  cross  in which the order and map position of the genes under study are not known. In this particular cross, the genes by which the parents differ will be  azi  (resistance or sensitivity to sodium azide),  gal  (ability or inability to utilize galactose as an energy source),  lac  (ability or inability to utilize lactose as an energy source), and  ton  (resistance or sensitivity to bacteriophage T1).  A  streptomycin-sensitivity  allele   in the Hfr and a streptomycin-resistance allele  (str r ) in the recipient are used to selectively kill the Hfr cells after  conjugation .

13. Continu: The relative positions of the  azi, ton, lac,  and  gal  genes were established in our experiment. However, the chromosomal region containing these loci might be only a small proportion of the entire  chromosome . The complete map is obtained from many such interrupted  conjugation  experiments, in which parental strains heteroallelic for different combinations of genes are used; then the overall map is pieced together from the complete set of data.

14. High-Resolution Mapping by Recombinant Frequency: Interrupted-mating experiments provide a rough set of  gene  locations over the entire map. As we learned, the genes are mapped by time of entry. In such experiments, the  exogenote  must integrate by a double  recombination  event, but the mapping method is not based on any measurement of  recombinant frequencies . However, to provide a higher-resolution method for measuring the sizes of  smaller map distances, recombinant frequencies are used.

15. Continu: Suppose that we undertake an experiment to map three genes— met, arg,  and  leu —by  recombinant frequency . To measure  recombination  between these genes, we must set up a  merozygote  that is heterozygous for all three. This can be accomplished if we can establish which  gene  enters last by an interrupted conjugation analysis. The Hfr  allele  of the last-entering gene is selected among the F − exconjugants.

16. Continu: The last  gene  to enter is  leu + ;  therefore we select initially for  leu +  exconjugants by plating them on  medium  containing no leucine but containing methionine and arginine. Now we can proceed to calculate map distance in the standard way by using a map unit equal to a  recombinant frequency  of 1 percent. In practice, this calculation is done by measuring the proportion of the total  leu +  exconjugants that also carries  arg +  or  met +  or both or neither. The  recombination  events needed to produce these recombinant genotypes are know that a  double crossover  must have occurred to integrate leu + :  one crossover is at the left of the  leu  gene, but the other can be in various positions at the right. 

17. Refernces:  Treanor, Brian,  Aspects of alterity: Levinas, Marcel, and the contemporary debate , Fordham University Press, 2006, p.41 Jump up^  Klein, Ernest,  A comprehensive etymological dictionary of the English language , Vol II, Elsevier publishing company, Amsterdam, 1969, p.1317 Jump up^   Saeed, John, Semantics, Blackwell, p. 12,  ISBN   0-631-22693-1