Genetics & Genotyping By Kristin Gill & Daisy Robinton HC70AL Spring 2009

What are the possible genotypes? Homozygous Wildtype Hemizygous Wildtype/T-DNA Homozygous T-DNA

What is the expected ratio of genotypes? 1 : 2 : 1 Homozygous Wildtype Hemizygous Homozygous T-DNA

What are the primer interactions? Wildtype Product = FORWARD + REVERSE T-DNA Product = LBb1 + FORWARD OR LBb1 + REVERSE

Structure of a Gene + Primers Given Primers from STOCKS Forward: 5’ ACTG 3’ Reverse: 5’ ATAA 3’ In Sequence Look For Forward: 5’ ACTG 3’ Reverse: 5’ TTAT 3’ FW 5’5’ 5’5’ 3’3’ 3’3’ A C T G T G A C 5’3’ A A T A T T A T RV 3’ 5’ Reverse Complement

What is a Knock-Out? T-DNA genotype based on FORWARD Primer & LBb1 Primer Interaction FW 5’5’ 5’5’ 3’3’ 3’3’ A C T G T G A C 5’3’ A A T A T T A T RV 3’ 5’ LBb1 5 kb T-DNA Insert

What is a Knock-Out? T-DNA genotype based on REVERSE Primer & LBb1 Primer Interaction FW 5’5’ 5’5’ 3’3’ 3’3’ A C T G T G A C 5’3’ A A T A T T A T RV 3’ 5’ LBb1 5 kb T-DNA Insert