UT BIO 325L - Homework12bSpring2014[KEY](1) (7 pages)

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Homework12bSpring2014[KEY](1)



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Homework12bSpring2014[KEY](1)

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Pages:
7
School:
University of Texas at Austin
Course:
Bio 325l - Laboratory Experience in Genetics

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Name Lab room day time TA name Arabidopsis 1 Controls are important for determining the validity of experimental results a 1 points For the genomic PCR which reactions were used as negative controls Positive controls Defend your answers All reactions using water instead of template were negative controls because they were expected in all cases to not produce a result The one reaction that may be less obviously a negative control is the reaction using the T DNA specific insert primers against the wild type plant This should not have produced a PCR band because there should not have been any T DNA in the wild type plants The PCR using genome specific primers against the wild type plant was the only positive control because the primers should not have any interrupting T DNA insertion between them and so should have generated a band b 0 5 points There is one more reaction that could have been used as a positive control with the genomic DNA Explain what this reaction should have The missing positive control would be one for a T DNA insert This would need the T DNA primers combined with a DNA template that definitively had an insert present 2 1 point What should the test PCR results be band or no band for the following situations Assume that T DNA inserts are disruptive to RNA expression a Genomic PCRs with gene specific and T DNA test primer pair sets on i A test plant lacking T DNA inserts ii A test plant heterozygous for a T DNA insert iii A test plant homozygous for T DNA inserts b RT PCR using a gene specific primer pair on i A test plant lacking T DNA inserts ii A test plant heterozygous for a T DNA insert iii A test plant homozygous for T DNA inserts Summary chart gDNA Gene specific primer pair T DNA primer pair test plant test plant No T DNA Heterozygote Homozygote for T DNA RT PCR Gene specific primer pair test plant 3 1 points Ideally we want to identify a plant that is homozygous for a disruptive T DNA insert If that does not happen we can self cross a



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