Genome Alterations and New Products of Biotechnology:- A Bacterial Plant Parasite Aids Cloning in Plants

We not only can understand genomes; we can change them. This is perhaps the ultimate manifestation of the new technologies. The introduction of recombinant DNA into plants has enormous implications for agriculture, making possible the alteration of the nutritional profile or yield of crops or their resistance to environ mental stresses, such as insect pests, diseases, cold, salinity, and drought. Fertile plants of some species may be generated from a single transformed cell, so that an introduced gene passes to progeny through the seeds. As yet, researchers have not found any naturally oc curring plant cell plasmids to facilitate cloning in plants, so the biggest technical challenge is getting DNA into plant cells. An important and adaptable ally in this ef fort is the soil bacterium Agrobacterium tumefaciens. This bacterium can invade plants at the site of a wound, transform nearby cells, and induce them to form a tu mor called a crown gall. Agrobacterium contains the large (200,000 bp) Ti plasmid (Fig. 9–26a). When the bacterium is in contact with a damaged plant cell, a 23,000 bp segment of the Ti plasmid called T DNA is transferred from the plasmid and integrated at a random position in one of the plant cell chromosomes (Fig. 9–26b). The transfer of T DNA from Agrobacterium to the plant cell chromosome depends on two 25 bp re peats that flank the T DNA and on the products of the virulence (vir) genes on the Ti plasmid (Fig. 9–26a). The T DNA encodes enzymes that convert plant metabolites to two classes of compounds that benefit the bacterium (Fig. 9–27). The first group consists of plant growth hormones (auxins and cytokinins) that stimulate growth of the transformed plant cells to form the crown gall tumor. The second constitutes a series of unusual amino acids called opines, which serve as a food source for the bacterium. The opines are produced in high concentrations in the tumor cells and secreted to the surroundings, where they can be metabolized only by Agrobacterium, using enzymes encoded elsewhere on the Ti plasmid. The bacterium thereby diverts plant resources by converting them to a form that benefits only itself.

FIGURE 9–26 Transfer of DNA to plant cells by a bacterial parasite. (a) The Ti (tumor-inducing) plasmid of Agrobacterium tumefaciens. (b) Wounded plant cells produce and release the phenolic compound acetosyringone. When Agrobacterium detects this compound, the virulence (vir) genes on the Ti plasmid are expressed. The vir genes encode enzymes needed to introduce the T DNA segment of the Ti plasmid into the genome of nearby plant cells. A single-stranded copy of the T DNA is synthesized and transferred to the plant cell, where it is converted to duplex DNA and integrated into a plant cell chromosome. The T DNA encodes enzymes that synthesize both plant growth hormones and opines (see Fig. 9–27); the latter compounds are metabolized (as a nutrient source) only by Agrobacterium. Ex pression of the T DNA genes by transformed plant cells thus leads to both aberrant plant cell growth (tumor formation) and the diversion of plant cell nutrients to the invading bacteria.

FIGURE 9–27 Metabolites produced in Agrobacterium-infected plant cells. Auxins and cytokinins are plant growth hormones. The most common auxin, indole acetate, is derived from tryptophan. Cytokinins are adenine derivatives. Opines generally are derived from amino acid precursors; at least 14 different opines are produced by enzymes en coded by the Ti plasmids of different Agrobacterium species.

This rare example of DNA transfer from a prokary ote to a eukaryotic cell is a natural genetic engineering process—one that researchers can harness to transfer recombinant DNA (instead of T DNA) to the plant genome. A common cloning strategy employs an Agrobacterium with two different recombinant plas mids. The first is a Ti plasmid from which the T DNA segment has been removed in the laboratory (Fig. 9–28a). The second is an Agrobacterium–E. coli shuttle vector in which the 25 bp repeats of the T DNA flank a foreign gene that the researcher wants to introduce into the plant cell, along with a selectable marker such as resistance to the antibiotic kanamycin (Fig. 9–28b). The engineered Agrobacterium is used to infect a leaf, but crown galls are not formed because the T DNA genes for the auxin, cytokinin, and opine biosynthetic enzymes are absent from both plasmids. Instead, the vir gene products from the altered Ti plasmid direct the trans formation of the plant cells by the foreign gene—the gene flanked by the T DNA 25 bp repeats in the second plasmid. The transformed plant cells can be selected by growth on agar plates that contain kanamycin, and addition of growth hormones induces the formation of new plants that contain the foreign gene in every cell. The successful transfer of recombinant DNA into plants was vividly illustrated by an experiment in which the luciferase gene from fireflies was introduced into the cells of a tobacco plant (Fig. 9–29)—a favorite plant for transformation experiments because its cells are particularly easy to transform with Agrobacterium. The potential of this technology is not limited to the pro duction of glow-in-the-dark plants, of course. The same approach has been used to produce crop plants that are resistant to herbicides, plant viruses, and insect pests (Fig. 9–30). Potential benefits include increased yields and less need for environmentally harmful agricultural chemicals.

Biotechnology can introduce new traits into a plant much faster than traditional methods of plant breeding. A prominent example is the development of soybeans that are resistant to the general herbicide glyphosate (the active ingredient in the product RoundUp). Glyphosate breaks down rapidly in the environment (glyphosate sensitive plants can be planted in a treated area after as little as 48 hours), and its use does not generally lead to contamination of groundwater or carryover from one year to the next. A field of glyphosate-resistant soybeans can be treated once with glyphosate during a summer grow ing season to eliminate essentially all weeds in the field, while leaving the soybeans unaffected (Fig. 9–31). Potential pitfalls of the technology, such as the evolution of glyphosate-resistant weeds or the escape of difficult to-control recombinant plants, remain a concern of re searchers and the public.

FIGURE 9–28 A two-plasmid strategy to create a recombinant plant. (a) One plasmid is a modified Ti plasmid that contains the vir genes but lacks T DNA. (b) The other plasmid contains a segment of DNA that bears both a foreign gene (the gene of interest, e.g., the gene for the insecticidal protein described in Fig. 9–30) and an antibiotic resistance element (here, kanamycin resistance), flanked by the two 25 bp repeats of T DNA that are required for transfer of the plasmid genes to the plant chromosome. This plasmid also contains the replication origin needed for propagation in Agrobacterium. When bacteria invade at the site of a wound (the edge of the cut leaf), the vir genes on the first plasmid mediate transfer into the plant genome of the segment of the second plasmid that is flanked by the 25 bp repeats. Leaf segments are placed on an agar dish that contains both kanamycin and appropriate levels of plant growth hormones, and new plants are generated from segments with the transformed cells. Nontrans formed cells are killed by the kanamycin. The foreign gene and the antibiotic-resistance element are normally transferred together, so plant cells that grow in this medium generally contain the foreign gene.

FIGURE 9–29 A tobacco plant expressing the gene for firefly luciferase. Light was produced after the plant was watered with a solution containing luciferin, the substrate for the light-producing luciferase enzyme (see Box 13–2). Don’t expect glow-in-the-dark ornamental plants at your local plant nursery anytime soon. The light is actually quite weak; this photograph required a 24-hour exposure. The real point—that this tech nology allows the introduction of new traits into plants—is nevertheless elegantly made.

FIGURE 9–30 Tomato plants engineered to be resistant to insect larvae. Two tomato plants were exposed to equal numbers of moth larvae. The plant on the left has not been genetically altered. The plant on the right expresses a gene for a protein toxin derived from the bacterium Bacillus thuringiensis. This protein, introduced by a protocol similar to that depicted in Figure 9–28, is toxic to the larvae of some moth species while being harmless to humans and other organisms. Insect resistance has also been genetically engineered in cotton and other plants.

اخر الاخبار

اخبار العتبة العباسية المقدسة

وفد العتبة العباسية المقدسة يزور شعبة الإعداد البدني للاطّلاع على برنامج تأهيل المنتسبين الجدد

قسم بين الحرمين يقدم جهودًا مضاعفة لإعادة المفقودات إلى الزائرين

بعد برامج تدريبية متخصصة.. متحف الكفيل يبيّن منجزات ملاكاته في فن الخاتم (الأرابيسك)

شعبة مدارس الكفيل الدينية تعقد الاجتماع التحضيري الأول لمهرجان روح النبوّة الثقافي

المزيد

الآخبار الصحية

ماذا يحدث إذا تناولنا الطعام مرة واحدة فقط في اليوم

اكتشاف عوامل خطر غير متوقعة لمرض الكبد الدهني

فيروس هانتا والسفينة الموبوءة.. تحديث من الصحة العالمية

طبيب يحسم الجدل السجائر الإلكترونية وعلاقتها بالسرطان

المزيد

الآخبار التكنلوجية

لأول مرة في الولايات المتحدة.. تشغيل مفاعل مصغر نووي يغذي الذكاء الاصطناعي بالطاقة

الكشف عن تفاصيل أول عملية ناجحة لاستعادة انتاج الحيوانات المنوية

أطلس يكشف خفايا الجسم البشري بدقة أرفع بـ50 مرة من شعر الإنسان

المتحف المصري الكبير يتحول إلى "أيقونة خضراء" بدعم ياباني

المزيد

مواضيع ذات صلة

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From Genomes to Proteomes:- Sequence or Structural Relationships Provide Information on Protein Function

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From Genes to Genomes:- DNA Libraries Provide Specialized Catalogs of Genetic Information

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