STRAINIMPROVEMENT NIKHIL AGGARWAL Roll No.- 09010620
DefinitionThe directed improvement of productformation or cellular properties throughmodifications of specific biochemicalpathways or by introduction of new pathwaysusing recombinant DNA technology.
The need• Heterologous protein formation• Extension of substrate range• Pathways leading to new products• Reduction of by-product formation• Improvement of yield etc.
Methods• Mutation• Modification of Gene Expression• Recombination
Mutation• Spontaneous mutations arise occasionally in all cells and develop in the absence of any added agent.• Induced mutations are the result of exposure of the organism to a mutagen.
Mutation contd..Mutagens:-• Base analogs- example 5-bromouracil (5-BU), an analog of thymine.• Intercalating agents- distort DNA to induce single nucleotide pair insertions and deletions. These include acridines such as proflavin and acridine orange.• Alkylating agents- causes changes in a base’s structure and therefore alters its base pairing characteristics. Example methyl- nitrosoguanidine, that adds methyl groups to guanine, causing it to mispair with thymine.• UV radiation, carcinogens such as aflatoxin B1 and other benzopyrene derivatives.
Modification of Gene Expression• It is possible to modify gene regulation by changing gene transcription, fusing proteins, creating hybrid promoters, and removing feedback regulation controls. These approaches make it possible to overproduce a wide variety of products.• It can also be done by altering controls or blocking enzymes in biochemical pathway.
Recombination• recombination in fungi: sexual and parasexual cycles• recombination in bacteria: transformation, transduction and conjugation• in vivo rearrangements by transposable genetic elements• protoplast fusion• in vitro recombinant DNA techniques
Applications• The enhancement of production of asperenone , an inhibitor of lipoxygenase and human platelet aggregation from Aspergillus niger CFTRI 1105, was achieved by UV and nitrous acid mutagenesis.• Nitrous acid mutants exhibited increased asperenone production when compared with UV irradiated mutants.
Applications contd..• Ethanol yeast normally grows at 25- 30°C and stop growing at 40°C while an optimum temperature for ethanol fermentation is 5-10°C higher.• Ethanol also has effect on microbial physiology. Its growth rate decreases when ethanol concentration is increased.• In order to enhance ethanol production yield in the fermentation, improving the yeast strain by two mutagenesis techniques as UV irradiation and chemical mutagen EMS were carried out.
Applications contd..• Firstly, Streptomyces albus JCM 4703 was sequentially treated with mutagens(UV, NTG, nitrogen mustard, etc.) to form strain SAM-X which produces 10mg/ml of salinomycin as compared to 250 μg/ml in JCM 4703.• Secondly, spontaneous mutants of SAM-X resistant to streptomycin, gentamicin, or rifampin were developed. Mutants with enhanced salinomycin production were detected from the Strr, Genr, or Rifr mutant isolates. The most productive strains were designated KO-600, KO-602, and KO-603, respectively.
ConclusionThe exploitation of cellular complexity for strainimprovement has been a challenging goal forapplied biological research. Progress in strainimprovement will depend not only on advancesin technologies for high-throughputmeasurements but, more importantly, on thedevelopment of theoretical methods thatincrease the information content of thesemeasurements and, as such, facilitate theelucidation of mechanisms and the identificationof genetic targets for modification.
Bibliography• www.wikipedia.org• http://www.scribd.com/doc/8801762/Isolation-Preservation-and- Improvement-of-Industrial-Microorganism• Microbiology by Lansing M. Prescott, 5th Ed.• Bioreaction Engineering Principles by John Villadsen• Research Papers:-1. Innovative Approach for Improvement of an Antibiotic-Overproducing Industrial Strain of Streptomyces albus (Norimasa Tamehiro, Takeshi Hosaka, Jun Xu, Haifeng Hu, Noboru Otake, Kozo Ochi)2. Strain improvement of Aspergillus niger for the enhanced production of asperenone (C. Chidananda , C. Mohan Kumar , A. P. Sattur)3. Strain Improvement of Ethanol Fermenting Yeast Using Random Mutagenesis Technique (Teerapatr Srinorakutara, Podjana Chumkhunthod, Suthkamol Suttikul, Wandee Yindeeyoungyeon, Ladda Wattanasiritham, Bancha Mouthung and Montree Wangpila)