Purification, concentration and recovery of small fragments of DNA from Giardia lamblia and their use for other molecular techniques

Abstract

Graphical abstract

Guidelines for Sanger sequencing and molecular assay monitoring

“…Examples include physical methods such as electrophoresis into a preformed trough, enzymatic methods such as agarose digestion, or purification methods using columns or magnetic beads. 5,18 Although in theory it is possible, given the specificity of the sequencing primers, to generate NA sequence information from a single amplicon included in a multiplex PCR reaction, this is an approach that is prone to primer interference, lower product yields, and increased analysis complexity, and therefore is not recommended for diagnostic applications. In most cases, repeating the PCR reaction using one specific pair of primers will resolve the problem.…”

“…Numerous options are available for this PCR product purification step, including bead-based, column-based, and enzymatic methods, all of which are available commercially. 5,18 For VDLs that outsource their sequencing, commercial sequencing laboratories typically provide, via their websites, a list of recommendations or preferences for specific clean-up kits and procedures to be used prior to submitting samples to their facility for sequencing. Many sequencing facilities also provide PCR purification services for an additional fee.…”