Hartl, Jones - Genetics. Principlers and analysis - 1998 (522927), страница 103

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The neo-r resistance marker is then used toselect plasmids containing the disease gene aftertransfer into E. coli, where the gene can beanalyzed more safely and easily.in Chapter 9.) The result is a heterogeneous collection of plasmids, most containing DNA fragments other than theone of interest. However, plasmids that do contain the fragment of interest also contain the transposon tag and soare able to confer antibiotic resistance on the host. The heterogeneous collection of plasmids is introduced into E.coli cells by transformation, and the cells are grown on solid medium containing the antibiotic. The only cells thatsurvive contain the desired plasmid, which contains the transposon along with DNA sequences from the desiredgene. The transposon-tagged gene is said to be cloned in E.

coli.The cloning of genes in E. coli greatly facilitates further genetic and molecular studies, such as DNA sequencing.The disease gene usually poses no hazard when it is present in E. coli for several reasons: (1) More than one genein the pathogen is usually required to cause disease; and (2) the cloned transposon-tagged gene is inactivated by theinsertion; and (3) the disease gene cloned in E. coli is usually incomplete or, if complete, is often inactive in thishost. In cases in which some danger might result from cloning, potential problems are minimized by using specialkinds of E. coli strains and following procedures that ensure containment in the laboratory.Chapter SummaryDNA can be transferred between bacteria in three ways: transformation, transduction, and conjugation.

Intransformation, free DNA molecules, obtained from donor cells, are taken up by recipient cells; by arecombinational mechanism, a single-stranded segment becomes integrated into the recipient chromosome,replacing a homologous segment.In conjugation, donor and recipient cells pair and a single strand of DNA is transferred by rolling-circle replicationfrom the donor cell to the recipient. Transfer is mediated by the transfer genes of the F plasmid. When F is presentas a free plasmid, it becomes established in the recipient as an autonomously replicating plasmid; if F is integratedinto the donor chromosome—that is,<b>Текст обрезан, так как является слишком большим</b>.

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