In order to genetically engineer a plant, one must be able to insert a gene into the genome of an individual plant cell and then cause that cell to differentiate into a whole plant. The former process is referred to as transformation; the latter, regeneration. The most common way of transforming cells exploits the ability of Agrobacterium tumefaciens, the causative agent of a common plant disease known as ‘crown gall.' Agrobacterium contains a closed-circular piece of double-stranded DNA called the tumor-inducing (Ti) plasmid. During infection, Agrobacterium inserts a segment of the Ti plasmid, called T-DNA (transferred DNA), into the plant's nuclear genome. This T-DNA contains genes encoding enzymes that catalyze the synthesis of plant growth regulators (cytokinin and auxin) which together control cell proliferation. This results in the formation of a tumor, within which the bacterium resides. The T-DNA also contains genes encoding enzymes that catalyze the synthesis of unique amino acids that the plant cannot utilize, and that serve as a carbon source for the bacterium.
[...] Platforms for Studying Tree Biology Marker-Aided Selection Marker-aided selection (MAS) involves the identiﬁcation of individuals based on the presence of DNA markers in offspring derived from parents whose genomes have already been mapped. DNA markers are usually random nucleotide sequences that do not encode a functional gene; they are frequently ampliﬁed via PCR and are visualized on a gel. The position of the markers on a chromosome is mapped by determining the frequency of their mutual recombination when haploid gametes are formed in a given individual. [...]
[...] Because the selectable marker gene is directly linked to the gene of interest (transgene), it too should be present in the transformed cell. Another DNA delivery system, biolistics, involves coating microscopic beads (usually gold or tungsten) with DNA. These beads are propelled at an explant, usually with a burst of compressed air as the driving force. Once inside the cell, DNA that sloughs off the bead can recombine with a plant chromosome. Biolistics is often less efﬁcient than Agrobacterium- mediated transformation because of cellular damage from the impact of the beads, digestion of the transgene by cytosolic enzymes (nucleases), and the need for recombination. [...]
[...] Although the potential beneﬁt of these traits has also been demonstrated in transgenic trees, no such trees are currently being grown in the USA for commercial purposes. Agronomic crops generally are herbaceous annuals that are highly domesticated and have few, if any, wild relatives. Most tree plantations are established in close proximity to wild or feral relatives, increasing the probability of transgene spread. Thus, a major concern over the use of transgenic trees is the potential for extensive transgene dispersal through pollen and seeds. [...]
[...] Conducting this work in a laboratory obviates the need for expensive ﬁeld trials, and forest managers will realize the additional genetic gain soon after crossing the selections from the previous generation. Sexual reproduction following MAS is possible, but gains will be fewer and greatly delayed relative to the clonal propagation. This is because juvenile selections must reach maturity, and the planting stock will not be genetically identical to the parents. However, wide-scale adoption of MAS for tree improvement has not been realized. [...]
[...] This pair of enzyme types allows molecular biologists to mix and match various coding sequences and promoters. Reporter genes allow one to visualize a promoter's expression pattern. The most common reporter gene, b-glucuronidase encodes an enzyme that catalyzes the conversion of a colorless substrate to an insoluble, blue-colored product that precipitates in cells expressing the gene encoding it. Polymerase chain reaction (PCR) is a technique that allows for ampliﬁcation of a speciﬁc piece of DNA. Short pieces of DNA (usually about 15 to 30 nucleotides in length), referred to as primers, are designed to complement sites on opposite strands of the target DNA. [...]
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