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Exercise 17 Bio 112 Genetics Lab

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There are four grain phenotypes in the above ear of corn: Purple & Starchy(A), Purple & Sweet(B), Yellow & Starchy(C) and Yellow & Sweet(D). These four grain phenotypes are produced by the following two pairs of heterozygous genes (R & r and SU & su) located on two pairs of homologous chromosomes (each gene on a separate chromosome): Dominant alleles Recessive alleles R = Purple r = Yellow SU = Starchy su = Shrunken The cross which produced the corn ear above was: Rr SUsu X Rr SUsu Dihybrid Cross in Corn

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DNA Extraction from Bacteria

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Transformation of Bacteria (pGLO)

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We will introduce a gene into E. coli that will produce a protein (green fluorescent protein-GFP) that will cause the colonies to glow green when exposed to ultraviolet light

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What is Transformation? Uptake of DNA (in this case a plasmid pGLO) from the surrounding environment of the cell.

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What is Green Fluorescent Protein- GFP)? GFP was discovered in the bioluminescent jelly Aequorea victoria. The gene that makes this protein is used extensively in researchand also for fun?

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12 microliters of DNA

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Transformation Procedure in a nutshell. Suspend bacterial colonies in Transformation Solution, CaCl 2 Add pGLO plasmid DNA to +DNA tube Place tubes on ice Heat shock at 42 o C and place on ice Incubate with LB broth Streak plates

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Expected Results

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Our genes of interest: amparaCGFP amp this gene will give our transgenic bacteria resistance to the antibiotic ampicillin araC this gene will produce a protein which in the presence of the sugar arabinose will allow the bacteria to turn on the GFP gene GFP in the presence of arabinose, this gene will turn on and cause the transformed (transgenic) bacteria to glow green pGLO ori bla GFP araC

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The Role of Arabinose The bacterial genes that make the digestive enzymes needed to break down arabinose for food are not expressed (made) when arabinose is absent. When arabinose is present the genes are turned on. When it is absent the genes remain off. Arabinose initiates transcription of the genes by promoting the binding of RNA polymerase to the promoter.

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pGLO ori bla GFP araC

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Expected Results