DONATIEN ISHIMWE 2 nd Presentation/DESIGNING References: 1)DNA isolation_www.arizona.edu 2)Cell isolation_www.plants.usda.gov. 3)Videos_www.youtube.com

My DESIGNINGQuestion No.1: HOW CAN WE CREAT THOSE NEW CASSAVA SPECIES? Answer: Genetic transformation of Cassava cell plant. Steps to get Cassava meristem cell 1)Cutting the apical meristem tissue into different parts And treat them with : Proteolytic enzymes like trypsin and collagenase to digest proteins in the extracellular matrix. Some reagents (Eg: ethylenediaminetetraacetic acid, or EDTA) that bind, or chelate, the Ca2+ on which cell-cell adhesion depends. As they form a suspension,the tissue can then be teased apart into single living cells by gentle agitation 2)Applying centrifugation. 3)Tissue Culture Aim: Clonage Conditions: Specific nutrients&Balanced nutrients-salts, vitamin,water and room temperature In order to have many cell to carry on the experiment,they put the small sample in a test tube They biologist call IN VITRO,so that they can make a clonage of the obtained single cell.

PROCESSES OF CASSAVA DNA EXTRACTION AND MODIFICATION Recombinant DNA technology to be used here will consist of INSERTION rather than removal. The series of procedures to be used to join together (recombine) DNA segments from other plants Genome that show the sequence of Nitrogenic bases that can be used by cell to produce proteins containing Valine as one of amino acid. Other possible recombinant of DNA molecule can constructed from segments of two or more different DNA molecules. Under certain conditions, like the breaking enzymes(endolygase>DNA helicases,..) a recombinant DNA molecule will enter a cell and replicate in new host cell nucleus, either on its own or after it has been integrated into a chromosome. DNA EXRTACTION : Seven steps of isolating DNA. 1)Grinding the tissue coffee grinder. 2)Making buffered solution with CTAB solution incubated at 60 0 C for 30mins. 3)Pouring chloroform into solution to separate DNA from protoplasm contents like lipids that soluble in chloroform rather than Water. 5)Centrifugation of 5mins in order to let the two formed suspensions separate each other. 6)Retrieving upper layer containing water and DNA will be puppetted. 7)A safe removal of whitish beards, the separated DNA. DNA INSERTION: This is a complicated step, at first DNA must be segmented by using the specific enzymes that break down the the hydrogen bonds joining the nitrogenic bases in the helix.

Results expectation of my Genetic modification: The new cassava plant species those can have protein and free of Linamarin (LFC-1 and LFC-2) As I am targeted to deviate the biosynthesis of Linamarin,the cause of CASSAVA INTOXICATION, in order to let its quantity being used in PROTEIN synthesis due to the presence mRNA with the anticodon that can be used to template the proteins with Valine as one of its amino acids. There fore Linamarin that would be produced,can be inhibited in that way,with an other additional feature of having Protein as the difference between those two new improved cassava species.

CLONAGE and Tissue Culture The modified cells would put in medium with specific nutrients and balanced salts contents in order to make them multiplied and make tissues we can allow to grow into plants. References: 1)DNA isolation_www.arizona.edu 2)Cell isolation_www.plants.usda.gov. 3)Videos_www.youtube.com