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GENETIC ENGINEERING.
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Use in agriculture, methods used. Example of genetically engineered rice.... More...
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Paper Abstract:
Use in agriculture, methods used. Example of genetically engineered rice.

Paper Introduction:
Genetic engineering is being used more and more in agriculture to produce plants which are resistant to disease and thus increase crop yields. This research paper will examine methods used to produce transgenic plants and an experiment which produced genetically engineered rice with resistance to sheath blight. Transgenic plants are produced for three major reasons. (Glick and Pasternack, 1998, p. 427). Firstly, transgenic plants often improve crop yields by inducing resistance to pests and other factors which affect the yield, or improving the ornamental value of plants by developing new colors, or strains resistance to certain climactic conditions. Secondly, transgenic plants can be used to produce medically and commercial important proteins and metabolites in large amounts under

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The group synthesized an adaptor which consisted of twocomplimentary oligonucleotides (5'-GATCCAACAATGAAGCT-3' and 5'-CTCATTGTTG-3'). K. Transgenic plants are produced for three major reasons. Transgenic plants are created by inserting DNA sequences which codefor particular genes into plasmids derived from the host plants.Transformation can also be accomplished by introducing cloned genes into asmall number of cells of a plant tissue from which whole plants can beformed (Glick and Pasternack, 1998, p. Estimates of the maximum number of chitinase genesinserted into the rice genome were obtained by counting the number ofautoradiographic bands unique to the transgenic plants in digests ofgenomic DNA with EcoRV. Chitinases are the PR proteinswhich have received the most study. Eight out of 22assayed plants exhibited functional chitinase transgene in at least some ofthe transformants (p. 688).The transformation vector used was PGL2 (CaMVCHl11), which contains a ricechitinase gene. Genetic engineering of rice for resistance tosheath blight. These are often obtainedfrom bacteria, and can be recognized by their reaction to antibiotics. Preliminary studies showed the effectswere still present after two generations, but these studies need to beextended to know if seeds from crops used for the next generation of plantswill retain the fungal resistance. These results suggest that this methodwill also be useful for introducing other new genes into Indica riceplants. The researchers successfully developed transgenic rice plants whichexpressed a rice chitinase gene constitutively. The researchers introduced a rice chitinase gene under thecontrol of CaMV 35S promoter into Chinsuah Boro II, and Indica variety ofrice (Lin et al, 1995, p. One problem not addressed by the study was the amount of chitinasegene activity necessary to afford maximal protection of the rice plantsfrom fungal infection. (1995). After two days in culture, apiece of agar containing the fungus was placed in the middle of a hill andthe sheaths were covered with parafilm. Most plants synthesize an assortment of proteins in response toinvasion by a pathogen or abiotic stress referred to as pathogenesis-related (PR) proteins (Lin et al, 1995). Chitinases hydrolyze the beta-1linkages of the N-acetyl-D-glucosamine polymer, chitin, which is a majorcomponent of many fungal cell walls, and may play a protective role againstfungal pathogens. (Glick andPasternack, 1998, p. Biol.,2 , pp. Rice plants with highlevels of chitinase exhibited increased resistance to infection by thesheath blight, R. The chitinase gene encoding for class 1 chitinase was used in thisexperiment. 619-629. The activity of the transgenic gene was still active through twogenerations, and some progeny had higher levels than the parental strains.The transgenic chitinase was expressed in leaves, sheathes, and roots ofuninfected plants. Two plantsessentially had no infection in the upper half of the plant, andparticularly in the flag leaf which contributes the most to grain filling,and thus the success of a crop. Rice plants are subject to fungal infection with R. Inactivation of chitinase genesin subsequent generations of tobacco plants has been shown (Hart, Fischer,Neuhaus and Meins, 1992). K., Datta, K., Soltanifar, N., Donn, G., & Potrykus, I.(1992). 69 ).. Protoplast were isolated from cultures of embryonic callus cells byincubation with an enzyme mixture (p. Marker genes are often usedalong with the gene to be inserted into a plant cell to aid in detectionand quantification of the transgenic process. Pp. 688). solani,which results in sheath blight Lin et al (1995) attempted to develop atransgenic strain of rice which would resist such infection byincorporating a gene for chitinase into the rice genome. Chitinaseactivity was detected in all plants that gave a positive reaction inwestern blot analysis. A possible way to improve the fungal resistance of rice plants tosheath rot is to explore other vectors as a means of inserting thechitinase gene into the rice DNA. solaniswas inoculated on a potato-dextrose plate. Plants expressing chitinase constitutively appeared tobe more resistant to R. (1994).Enhanced protection against fungal attack by constitutive co-expression ofchitinase and gluconase genes in transgenic tobacco. 686-691. Different vectors may result in a higherproduction of chitinase in the transgenic plants, and thus a higherresistance to the fungus. 179-188. 13,. 688). Genetic engineering is being used more and more in agriculture toproduce plants which are resistant to disease and thus increase cropyields. The plants were bagged before anthesis and seedswere collected and stored in a cold room. In some transgenic plants, the chitinase levels were comparable tothat seen in infected plants that are maximally induced for chitinase (p.689). M., & Meins, F. Biotech. Hart, C. The numbers seen in this experiment ranged fromtwo to five (p. Although lesions appeared within three to four daysin both plants, the number and size of lesions in transgenic plants werewere smaller than controls (p. Only a limited number of plants were produced inthe trial study, and large-scale studies will be needed to determine theeffectiveness of these transgenic plants in producing crops with asufficient resistance to sheath rot to be of benefit commercially. Lin, W., Anuratha, C. Gen. The chitinasegene was chosen because previous studies have indicated that constitutiveexpression of chitinase in transgenic tobacco plants was associated withgreater resistance to infection by this fungus (Zhu, Maher, Masoud, Dixonand Lamb, 1994), and purified rice chitinase has been shown to inhibit thegrowth of the fungus in vitro. References Datta, S. 434). S., Datta, K., Potrykus, I., Muthukrishnan, S.,& Datta, S. Herbicide-resistant Indica rice plants from IRRI breeding lineIR72 after PEG-mediated transformation of protoplasts. Progeny from two transgenic plants which expressed chitinase, one at ahigh level and one at a low level, were assayed for resistance to attack bythe fungal pathogen. Plant. Firstly, transgenic plants often improve cropyields by inducing resistance to pests and other factors which affect theyield, or improving the ornamental value of plants by developing newcolors, or strains resistance to certain climactic conditions. Polyacrylamidegel electrophoresis and western blotting were performed to detect chitinaseactivity. Different vectors may also affect the passage ofthe chitinase gene to future generations. Mol. Mol. (1992).Regulated inactivation of homologous gene expression in transgenicNicotiana sylvestris plants containing a defense-related tobacco chitinasegene. This research paper will examine methods used to producetransgenic plants and an experiment which produced genetically engineeredrice with resistance to sheath blight. Hygromycin was added to theculture medium on day 14, and the level of hygromycin was maintained forfour weeks to select resistant cells. Genet., 235, pp. Genomic DNA was isolated from freeze dried rice leaves by thecetyltrimethylammonium bromide (CTAB) nucleic acid extraction procedure.In a bioassay, a sclerotium of the rice sheath blight pathogen R. Preliminary screening of transgenic plants for hygromycin resistanceshowed that most of the plants exhibited HPT activity. Transgenic plants also speed up the process bywhich new species or variants can be produced and can result in plants withnovel features. A., Masoud, S., Dixon, R. Regenerants were transferred to agreenhouse for maturity. Secondly,transgenic plants can be used to produce medically and commercial importantproteins and metabolites in large amounts under controlled conditions andwith reproducible attributes. For analysis, fresh or frozen tissue from leaves, sheaths, or roots ofcontrol and transgenic plants were extracted and hygromycinphosphotransferase (HPT) activity assays were performed. In the transgenic plants, the lesionswere confined to the lower half of the sheaf, but in the control plants,the lesions had spread to the upper half of the plant. Bio/Tech., 12, pp.8 7-812. The aboveexperiment clearly showed the beneficial effects of high level chitinaseactivity in protecting rice plants against sheath rot. M., Fischer, B., Neuhaus, J. Analysis of proteins from leaves of T1generation plants derived from primary transformants showed the chitinaseand HPT genes were also expressed in these plants. Theadaptor was used to connect the 3'end of CaMV promoter to the 5'end of theCHl11 rice chitinase gene. The protoplast suspension wasthen incubated with plasmid DNA and carrier calf thymus DNA.Polyethyleneglycol solution was then added, and the mixture incubated at2 oC for 1 min. This adaptor contains a plant translation start consensus sequence, aBamHl restriction site at the 5'-end and a Sacl site at the 3'-end. solani (p. 687). Further culturing was according to the method of Datta,Datta, Soltanifar, Donn and Potrykus (1992). Zhu, Q., Maher, E. A., & Lamb, C. Selected embryonic cells were pickedafter 1 to 14 weeks, transferred to regeneration medium and incubated inthe light for plant regeneration. 687). Southern blot analysiswas performed on DNA from the leaves of transgenic plants. Thirdly, genetic transformation provide ameans for studying the action of genes during various biological anddevelopmental processes. Studiesalso need to be carried out to determine if the protection is carried onthrough succeeding generations. solani as shown by bioassay. 427). The infected rice plants were keptin a greenhouse to maintain high humidity and symptoms caused by fungalinfection were scored at weekly intervals. Many of the transformed plants werefertile, and about a third of them expressed the unselected chitinase gene,while some had substantially high levels of expression.

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