Transgenic plants

Genetic Engineering and Biotechnology 4.  Gene Transfer Methods and Transgenic Organisms
Gene transfer methods 
Gene transfer (transfection) methods in animals
Gene transfer methods in plants
Transgenic Organisms
Transgenic animals 
Transgenic plants 
Transgenic plants
Transgenic plants have been produced in more than 50 species by the end of the year 1992 (Table 42.2) and are being produced in a number of additional species every year. The progress has been so spectacular that by the turn of the century, we hope to be growing crops, which have been tailored to market specification by addition, subtraction or modification of genes. Transgenic plants have been produced, which are resistant to herbicides, insects, viruses and a variety of other biotic and abiotic stresses.

Table 42.2.A list of higher plants where transgenic plants have been produced, using different methods.
I. Herbaceous dicotyledons
  1. Nicotiana tabacum (tobacco)
  2. N. plumbaginifolia (wild tobacco)
  3. Petunia hybrida (petunia)
  4. Lycopersicon esculentum (tomato)
  5. Solanum tuberosum (potato)
  6. Solanum melongena (eggplant)
  7. Arabidopsis thaliana
  8. Lactuca sativa (lettuce)
  9. Apium graveolens (celery)
  10. Helianthus annuus (sunflower)
  11. Linum usitatissimun (flax)
  12. Brassica napus (oilseed rape; canola)
  13. Brassica oleracea (cauliflower)
  14. Brassica oleracea var. capitata (cabbage)
  15. Brassica rapa syn. B. campestris
  16. Gossypium hirsutum (cotton)
  17. Beta vulgaris (sugarbeet)
  18. Glycine max (soybean)
  19. Pisum sativum (pea)
  20. Medicago sativa (alfalfa)
  21. M.varia
  22. Lotus corniculatum (lotus)
  23. Vigna aconitifolia
  24. Cucumis sativus (cucumber)
  25. Cucumis melo (muskmelon)
  26. Cichorium intybus (chicory)
  27. Daucus carota (carrot)
  28. Armoracia sp. (horse radish)
  29. Glycorrhiza glabra (licorice)
  30. Digitalis purpurea (foxglove)
  31. Ipomoea batatas (sweet potato)
  32. Ipomoea purpurea (morning glory)
  33. Fragaria sp. (strawberry)
  34. Actinidia chinensis (kiwi)
  35. Carica papaya (papaya)
  36. Vitis vinifera (grape)
  37. Vaccinium macrocarpon (cranberry)
  38. Dianthus caryophyllus (carnation)
  39. Chrysanthemam sp. (chrysanthemum)
  40. Rosa sp. (rose)
II. Woody dicotyledons
  1. Populus sp. (poplar)
  2. Malus sylvestris (apple)
  3. Pyrus communis (pear)
  4. Azadirachta indica (neem)
  5. Juglans regia (walnut)
III. Monocotyledons

  1. Asparagus sp. (asparagus)
  2. Dactylis glomerata (orchard grass)
  3. Secale cereale (rye)
  4. Oryza sativa (rice)
  5. Zea mays (corn)
  6. Triticum aestivum (wheat)
  7. Avena sativa (oats)
  8. Festuca arundinacea (lall fescue)
IV. Gymnosperms (a conifer)

  1. Picea glauca (white spruce)

Transgenic plants with herbicide resistance.
Due to increasing concern about contamination of environment due to herbicides, new herbicides are being developed that are safer and biodegradable. This has necessitated the development of resistance in crop plants against these new and safer herbicides. These newer herbicides affect processes like photosynthesis or biosynthesis of essential amino acids (Table 42.3). Transgenic plants resistant to these herbicides have been produced, utilizing one of the following two approaches : (i) Either the target protein in overexpressed (e.g. EPSPS) or it should become insensitive to the herbicide (e.g. ALS). (ii) A pathway is introduced, which will detoxify the herbicide. For instance, glutathione-S transferase (GST) detoxifies atrazine, 'nitrilase' coded by gene bxn, detoxifies 'bromoxynil' and 'phosphinothricin acetyl transferase (PAT)' coded by bar gene, detoxifies 'L-phosphinothricin (PPT)'. (For more details, see Table 42.3).

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Transgenic plants with insect resistance.
Genes for insect resistance from three different sources are being transferred for developing insect resistance (i) bt2 gene encoding Bt toxin, derived from Bacillus thuringiensis; (ii) cowpea trypsin inhibitor gene (CpTi) from cowpea (Vigna unguiculata)and (iii) genes for other insecticidal secondary metabolites derived from a number of legumes. In all these cases, successful attempts to transfer genes for insect resistance have already been made. Insect resistant transgenic plants derived thus are being field tested and may be released for commercial cultivation.

Transgenic plants with resistance against viruses. Three different approaches were used for developing virus resistant transgenic plants : (i) Genes for virus coat protein or capsid protein (CP) were transferred to tomato, alfalfa, tobacco, potato, melon and rice for developing resistance against a variety of viruses (e.g. TMV, PVX and PVY, etc.). (ii) Gene for nucleocapsid protein (N) was transferred from tomato spotted wilt virus (TSWV) and provided resistance against this virus, which causes considerable damage to crops like tomato, tobacco, groundnut, pepper, etc. (iii) DNA fragments coding for satellite RNAs (also called virusoids) associated with viruses (e.g. 'cucumber mosaic virus' or CMV and 'tobacco ringspot virus' or TobRV) were transferred and provided resistance against these and other viruses.

Transgenic plants with resistance against bacterial and fungal pathogens. Several examples are now available, where genes imparting resistance against bacterial and fungal pathogens have been successfully transferred to tomato (against Pseudomonas, Alternaria and Rhizoctonia)and potato (against Phytophthora).

Transgenic tomato plants for hard skin and improved flavour. In tomato using antisense RNA technology, transgenic plants have been produced which are either 'bruise resistant' (suitable for transport and storage) or exhibit 'delayed ripening' giving more time for ripening on the plant, thus permitting more time for sugar accumulation and also giving higher shelf life. These are given the name 'Flavr Savr'.

Transgenic plants for hybrid seed production. During 1990-92, genes barnase (encoding a ribonuclease) and barstar (encoding a ribonuclease inhibitor) derived from Bacillus amyloliquefaciens were separately transferred to Brassica napus to produce 'male sterile' and 'fertility restorer' plants. The gene constructs were prepared where barnase/barstar was fused with TA29 promoter form tobacco, permitting expression only in the anther causing male sterility or restoration. This system will prove to oe of immense value for hybrid seed production in crop plants in future.

Transgenic plants for molecular farming.
Transgenic plants are also proposed to be used as factories or bioreactors for manufacturing speciality chemicals or Pharmaceuticals. Sugars, fatty acids, starch, cellulose, rubber, wax, etc. are obtained from plants and transgenic plants can be produced to increase their production. Following are some examples : (i) Transgenic tobacco plants with increased level of mannitol were produced using a gene for mannitol dehydrogenase (this also gave resistance against salinity), (ii) Transgenic potato plants with increased level of cyclodextrins or CDs were produced using a bacterial gene for cyclodextrin glucosyl transferase or CGTase. (CDs are useful for pharmaceutical delivery systems, flavour and odour enhancement and for removing undesirable compounds like caffeine from food), (iii) Transgenic Arabidopsis plants for production of pplyhydroxybutyrate or PHB (a biodegradable thermoplastic polymer) were produced using two genes, phbB and phbC. (iv) Transgenic potato and tobacco plants for production of human serum albumin or HSA (in leaf tissue) could be successfully produced.

Transgenic plants for study of regulated gene expression. Transgenic plants have also been produced for identifying, regulatory sequences involved in the differential expression of gene activity in time and space or due to a specific stimulus like light, temperature, wound, etc. For this purpose, different lengths of upstream flanking sequences of a structural gene are fused with their own structural gene or with other unrelated structural genes studied in the transgenic plants thus produced. This allowed the identification of regulatory elements e.g. light response elements (LRE), endosperm box, heat shock element (HSE), etc. For instance when LRE was fused with an unrelated gene, the latter started behaving like a photosynthetic gene (expression in leaf and light, not in roots and dark). A number of genes, thus examined, are listed in Table 42.4.

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