2.2.3. Other relevant experiments
Recovery and re-amplification of polymorphic fragments: the
polymorphic DNA fragments were recovered from polyacrylamide gel.
The target band was cut with clean blade and put into centrifuge tube.
50 μL deionized water was added to wash the ion and other
impurities adsorbed on the gel surface. After washing, the water was
sucked out, and 20 μL deionized water was re-added and then boiled
for 10 min to release DNA. After cooling, the sample was centrifuged
for 5 min at 12,000 rpm, and the target fragment in supernatant was
re-amplified after centrifugation. The reaction system and thermal
cycling program of secondary amplification have been the same as
MSAP pre-amplification conditions. (The conditions and reaction
systems of restriction, adapter-ligation, pre-amplification and selective
amplification refer to Supplementary material 1).