Lack of independent replicates: A common pitfall in experimental design.

Replication the number of individual samples included in an average greater replication provides greater statistical strength –reduces variance; strengthens the generality of the mean

Replicates must be Independent all statistical analyses assume replicate values are independent observations –a single value does not depend on one, more than it does on another failures of the assumption of independence –lack of truly random sampling or assignment to treatment groups –pseudoreplication (Hurlbert, S.H Ecological Monographs 54(2): )

Inhibition of Biofilm Formation Chemical 1 Chemical 2 Chemical 3 Control

Replicates are not independent because they are all in the same dish Differences between dishes will be misinterpreted as differences between treatments

Inhibition of Biofilm Formation Chemical 1 Chemical 2 Chemical 3 Control

Pseudoreplication Stuart Hurlbert Ecological Monographs 54(2): Treating multiple measurements from/on the same sample the same as single measurements made from/on multiple samples The multiple measurements are not independent because they are all from the same sample; hence they are not replicates

At which temperature will Serratia marcescens grow best? 20  C25  C35  C45  C Measure absorbance of five samples from each flask.

Pseudoreplication because... Multiple measurements from the same flask are treated as if they were single measurements from multiple flasks Each flask represents a single replicate, because multiple samples taken from each flask are not independent of one another

Five flasks incubated at each temperature. 20  C25  C35  C45  C Measure absorbance of one sample from each of the 20 flasks.