For SDS-PAGE and Western blot, whole bacterial cell extracts or flagella extracts were denatured and separated in 14% polyacrylamide gels
and transferred onto polyvinylidene difluoride (PVDF) membranes (Millipore Corp., Bedford, MA) (27).
Isoelectric focusing was on linear, wide-range immobilized polyacrylamide gel
strips (17 cm; pH 3-10; Bio-Rad Laboratories) in the first dimension and on 15% polyacrylamide gels
(20 cm x 22 cm x 1 mm) in the second dimension.
Eight percent polyacrylamide gels
(Invitrogen) were also suitable in cases where primers C and D were used for the determination of the Hp 2 allele instead of primers A and B.
The digests were fractionated on nondenaturing 10% polyacrylamide gel
. The Bst EII pattern generated during PRA yielded two 235/210-bp bands similar to the patterns attributed to M.
DGGE was carried out essentially as described previously (12) with 20 [micro]L of sample loaded onto a 10% polyacrylamide gel
containing a 40-70% denaturing gradient (exon 2b); 8% polyacrylamide gel
containing a 25-65% or a 40-70% denaturing gradient (exons 3, 4, 5, and 6); 12% polyacrylamide gradient gel containing a 40-80% denaturing gradient (exon 2a, 3'-UTR).
Catalysts' for polyacrylamide gel
polymerization and detection of proteins by silver staining.
DNA bands on the polyacrylamide gel
were visualized by silver staining (Bio-Rad).
The different conformers are separated on a nondenaturing polyacrylamide gel
and detected either radioactively or with silver staining.
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A sodium dodecyl sulphate polyacrylamide gel
electrophoresis profile demonstrated that the [alpha]' and [alpha] subunits of [beta]-conglycinin and the acidic chains of glycinin gradually degraded.
However the method is performed and compared for quality mostly in polyacrylamide gel
to detect denatured Single stranded DNA9,10.