Arabidopsis Genetics By: Tyler Ashby, Lani Huynh, Laura Johnson, Susannah Robichaux

HYPOTHESIS We predicted that the genes responsible for the Anthocyanin pigment (PAP) and the trichome number (RTN) of the Recombinant Inbred Lines (RIL) would be continuous traits while the erecta phenotype would be discrete.

SEED PREPARATION A microfuge tube was filled halfway with dilute bleach solution 5 seeds from the same RIL were put into the microfuge tube with a solution of dilute bleach to soak for one minute The seeds were then rinsed five times with sterile water The sterilized seeds were then put in the corresponding petri dish 5 more seeds with from the same RIL underwent the sterilization, then 10 from each line

PETRI DISH PREPARATION & SET UP Clean petri dishes were labeled with the number of the RIL and our group name using a wax pencil - One petri dish was used for each RIL A small square of paper towel was wet with sterile water and placed in the bottom of each petri dish Ten sanitary seeds of the same RIL were placed in the corresponding plates The dishes were stored first in the refrigerator, as part of the cold shock treatment, and then in stacks of three in the light box and covered with Saran Wrap

RESULTS FROM PETRI DISHES The leaves of the lines that did sprout were too small to use for an accurate trichome count, but we counted them for practice. The sucrose solution had no effect on the color already dying plants, so we were unable to determine whether the anthocyanin pigmentation is a continuous trait or a discrete trait. The plants did not grow enough to determine the erecta phenotype so we could not tell whether the gene is continuous or discrete.

POTTED PLANTS 5 sterilized seeds were placed on one of two Jiffy pots in the cardboard pots for each line They were then watered with miracle grow solution until the soil was dark brown every class period Once the plants bore flowers, a protective shield made of laminate stopped the flowers from cross-pollinating

RESULTS The plants grew much more quickly-within a week there were shoots and leaves from every RIL. We were able to get a trichome count from the first true leaves of each line. The erecta phenotypes of the grown plants were noted on almost all lines.

TRICHOME COUNT PER LINE

NUMBER OF LINES WITH A SPECIFIC NUMBER OF TRICHOMES

ERECTA PHENOTYPE vs. NOT ERECTA PHENOTYPE

TESTING FOR ANTHOCYANIN PIGMENT Because the potted plants’ pigmentation could not be observed if the plants were directly watered with the sucrose solution, cuttings of the mature leaves were placed in drops of the sterile sucrose solution in petri dishes. – Two from each RIL This method did not work because the solution’s adhesive properties pulled the leaves into the drop. This would kill the leaves, so the pigmentation could not be measured. Because this didn’t work, a few drops of the sucrose solution were put into the bottom small microfuge tubes. The leaves were put, cut side down, into the solution. These were left overnight to grow and, hopefully, turn purple.

RESULTS The cuttings we put into the microfuge slid down into the sucrose solution and withered. There was no purple pigmentation and therefore no helpful data. Though we were unable to draw a conclusion about the anthocyanin pigmentation using our own data, the class’s data suggested it was a continuous trait. The number of trichomes was also a continuous trait, as seen by the varying number of counts for different lines. The erecta phenotype was discrete. This was observed rather than measured.