Analyze The Data

As genes encoding cyclin-dependent kinases were cloned from fission yeast, budding yeast, and animal cells, investigators working in plant systems designed experiments to determine whether plants also possessed CDKs that functioned as the catalysts for cell cycle progression. To isolate a cDNA encoding a CDK in maize, investigators aligned the predicted amino acid sequences of human cdc2, ,S. pombe cdc2, and sS. cerevisiae cdc28 genes (see the figure below). Degenerate

----menyqkvekigegtygvvyk

----medytkiekigegtygvvyk msgelanykrlekvgegtygvvyk arhklsg---rivamkkirledesegvpstaireis grhkttg---qvvamkkirleseeegvpstaireis aldlrpgqgqrvvalkkirlesedegvpstaireis

llkevndennrsncvrlldilhaes-klylvfefldmdlkkymdrisetgatsldprlvq llkelr----hpnivslqdvlmqds-rlylifeflsmdlkkyldsippg — qymdsslvk llkelk----ddnivrlydivhsdahklylvfefldldlkrymegipkd — qplgadivk

kftyqlvngvnfchsrriihrdlkpqn sylyqilqgivfchsrrvlhrdlkpqn kfmmqlckgiaychshrilhrdlkpon llidkegnlkladfglarsfgvplrnytheivt lliddkgtikladfglarafgipirvythevvt llinkdgnlklgdfglarafgvplraytheivt

lwyrapevllgsrhystgvdiwsvgcifaemirrsplfpgdseideifkifqvlgtpnee lwyrspevllgsarystpvdiwsigtifaelatkkplfhgdseidqlfrifralgtpnne lwyrapevllggkqystgvdtwsigcifaemcnrkpifsgdseidqifkifrvlgtpnea

vwpgvtllqdykstfprwkrmdlhkvvpngeedaiellsamlvydpahrisakralqqny vwpeveslqdykntfpkwkpgslashvknldengldllskmliydpakrisgkmalnhpy iwpdivylpdfkpsfpqwrrkdlsqvvpsldprgidlldkllaydpinrisarraaihpy .** . * *.* .**.*. * * . ..** . * *** .***. * . *

S. pombe cdc2 human cdc2 S. cerevisiae cdc28

S. pombe cdc2 human cdc2 S. cerevisiae cdc28

S. pombe cdc2 human cdc2 S. cerevisiae cdc28

S. pombe cdc2 human cdc2 S. cerevisiae cdc28

S. pombe cdc2 human cdc2 S. cerevisiae cdc28

S. pombe cdc2 human cdc2 S. cerevisiae cdc28

oligonucleotide PCR primers corresponding to the boxed amino acids were generated and a DNA fragment of the expected size was synthesized by PCR using these primers and maize cDNA library as a template. Why were the boxed regions selected as the basis for primer design?

The PCR product was then used as a probe to screen the maize cDNA library, and a full-length cDNA clone was isolated. The clone was sequenced, and the predicted amino acid sequence was 64% identical to human cdc2 and 63% identical to S. pombe cdc2 and to S. cerevisiae cdc28. What does the level of sequence similarity suggest about the evolution of cdc2 genes?

A complementation experiment was performed with sS. cerevisiae cells possessing a temperature-sensitive cdc28 mutation. Wild-type cells, cdc28ts cells, and cdc28ts cells transformed with the maize cdc2 cDNA under the influence of a strong promoter were grown at the permissive (25 °C) or restrictive (37 °C) temperatures. Cell proliferation was monitored by the growth of colonies on the culture plates (shown in the figure below).

37 °C

Why do the cdc28ts cells form colonies at 25 °C but not 37 °C? What is the significance of colony formation of the cdc28ts + maize cdc2 cells at 37 °C? What does this experiment tell us about the functional homology of cyclin-dependent kinase genes among eukaryotic species?

Your Heart and Nutrition

Your Heart and Nutrition

Prevention is better than a cure. Learn how to cherish your heart by taking the necessary means to keep it pumping healthily and steadily through your life.

Get My Free Ebook


Post a comment