Large Fragment Recovery

Objective:

Determine accuracy and recovery yields for size selection of large fragments using the Ranger

Accuracy Experiment:

Input genomic DNA was enzymatically fragmented to produce a distribution of DNA fragments that ranged in size from ~1 kbp to over 17 kbp. Roughly 2 ug of this input DNA was loaded onto the Ranger, which targeted recovery of DNA from 9 kbp to 13 kbp. Pre and post size selection fractions were characterized by a Bioanalyzer 2100 (Agilent Technologies).

Trace of the large fragment input DNA sample prior to size selection on the Ranger. Note that the Bioanalyzer marker does not constitute any part of the processed sample.

Trace of the large fragment input DNA sample prior to size selection on the Ranger. Note that the Bioanalyzer marker does not constitute any part of the processed sample.

 

Trace of Ranger output. Data points near full width-half max of peak height closely match the desired targets (9 kbp and 13 kbp). Note that the Bioanalyzer marker does not constitute any part of the processed sample.

Trace of Ranger output. Data points near full width-half max of peak height closely match the desired targets (9 kbp and 13 kbp). Note that the Bioanalyzer marker does not constitute any part of the processed sample.

Recovery Experiment:

Four replicate 20 ul aliquots were created for each of 10 kb and 20 kb NoLimit DNA fragments (Thermo Scientific, # SM1751, # SM1541). Replicates #3 and #4 from each set were run on the Ranger, which targeted the entirety of each band for recovery. The Ranger-processed replicates were concentrated, eluted in 17uL, and all four replicates were quantified using a Qubit Fluorometer (Life Technologies) to characterize recovery of these large fragments.

 

Concentration recovery data for size selection of 10 kbp and 20 kbp NoLimit fragments. All data generated with a Qubit Fluorometer.

Concentration recovery data for size selection of 10 kbp and 20 kbp NoLimit fragments. All data generated with a Qubit Fluorometer.