Molecular weight markers electrophoresis

Molecular weight markers for electrophoresis and hybridization techniques are widely used. These markers provide information in regard to molecular weights of rearranged bands. These techniques are useful in monitoring patients for relapse or residual disease status. The 32P-labeled and biotinylated DNA molecular weight markers provide visualization on the film and membrane. Hardware systems with band size computation capabilities are available. 

Fig. 30.2. Agarose gel electrophoresis of PCR product (lane 2) obtained from S. enterica serovar Typhimurium ATCC 15038. Lane 1 Molecular weight marker (<1>X174-Hinf I genome). No band was obtained with sterile water used as a negative PCR control (lane 3).

Compton, M. M., Lapp, S. A., and Pedemonte, R. (2002) Generation of multicoloured, prestained molecular weight markers for gel electrophoresis. Electrophoresis 23, 3262-3265. 

Fig. 1. Sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis (PAGE) analysis of expression and purification of recombinant protein. Ten-microliter aliquots were withdrawn at each step of the purification and loaded on a 12% SDS-PAGE gel in a Mini Protean III cell gel electrophoresis unit (Bio-Rad). The detection was performed with Coomassie blue staining. MW, low range (14-98 kDa) molecular weight marker (Bio-Rad).

The molecular weight markers should fall within established migration time ranges for the analysis to be acceptable. If the markers are outside this range, the gel electrophoresis run must be repeated. 

Fig. 6.14. Agarose gel electrophoresis of Echinococcus granulosus (horse strain) RNA. The samples in lanes 1,2,7 and 16 were prepared in 10 mM phosphate buffer, pH 6.8, and were not denatured. The samples in lanes 3, 4, 8 and 13-15 were denatured in 50% (w/v) dimethylsulphoxide in 10 mM phosphate buffer, pH 6.8. The samples in lanes 5,6,9 and 10 12 were denatured in 1 m glyoxal/3 m urea in 10mM phosphate buffer, pH 6.8. Lanes 1,3 and 5, E. granulosus total nucleic acids (10 fig) lanes 2,4 and 6, E. granulosus RNA (5 fig) lanes 7,8 and 9, Escherichia coli RNA (10 fig) lanes 10-16, Schistosoma mansoni RNA (10 fig). The E. coli RNA was included as a molecular weight marker the larger subunit is approximately 1 000 000 and the smaller subunit is 500 000. The RNA was visualised by staining with ethidium bromide and ultraviolet illumination. (After McManus et al., 1985.)...

Figure 11.1 Native polyacrylamide gel (5%) electrophoresis of purified fructosyltransferase. (a) Lane 1, molecular weight markers (45 kDa ovalbumin, 67kDa bovine serum albumin (BSA), and 134kDa BSA dimer) lane 2, pure fructosyltransferase. (b) Pure fructosyltransferase (one lane from the same gel was cut and stained for fructosyltransferase activity).

Figure 1. Apparent heterogeneity of humanized antibody hAB-1. Ten micrograms of each antibody were mixed with non-reducing or reducing SDS-PAGE sample buffer and boiled for ten minutes prior to electrophoresis. The NOVEX molecular weight markers (MW) are identified by their mass values. Samples Lanes 1,5 pMAB, lanes 2,6 hAB-2 and lanes 3,7 hAB-1. HC and LC correspond to heavy and light chains of antibodies, respectively.

Reproducibility of electrophoresis and Southern blotting is essential and can be maintained by careful quality control tests of the pH and conductivity of key reagents (e.g. electrophoresis buffer and SSC). The quality of fingerprints will be affected by poor Southern transfer of DNA. Therefore, transfer should always be checked by restaining blotted gels in electrophoresis buffer plus ethidium bromide (0.5 mg H) for 30 min. Inspection of the restained gel on a UV-transil-luminator should reveal no signs of residual DNA molecular weight markers. 

Fig. 3. Autoradiogram of P-ADP-ribosylation of Rho in Swiss 3T3 cell homogenate. Swiss 3T3 cells were treated with buffer alone (lane 1), 30 ng/ml of the wild type (lane 2) or the E173Q mutant (lane 3) of C3 exoenzyme for 72 h in DMEM containing 10 % fetal bovine serum. The cells were washed twice in phosphate-buffered saline (PBS), and incubated with 0.05 % (w/v) trypsin in PBS. The detached cells were collected and suspended in ADP-ribosylation buffer containing [ P]-NAD, followed by sonication. The homogenates were incubated with 100 ng of the wild type C3 enzyme at 30°C for 2 h. The reaction was terminated by the addition of trichloroacetic acid and Na deoxycholate. The pellets were subjected to 12 % SDS polyacrylamide gel electrophoresis and autoradiography. The positions of molecular weight markers are indicated on the left. The position of ADP-ribosylated Rho is indicated by an arrow...

Figure 3. 2-D Gel Electrophoresis of in vitro Translated Soybean Hypocotyl mRNA. Numbers to the left indicate migration of molecular weight markers (kilodaltons). The separating gel was 12% polyacrylamide. Numbered arrows indicate polypeptides that are up-regulated by BR while lettered arrows show polypeptides that are down-regulated in response to BR. Other experimental details are described in the text. A = hypocotyl sections auxin-depleted for 2 hours followed by buffer treatment for 2 hours A = as in A with 340 nM BR replacing buffer treatment.

Fig. 1. Two-dimensional NEPHGE/SDS-PAGE Gel of GT-labeled mouse liver nuclei. 5 xg of total mouse liver nuclear proteins were labeled with excess GT and UDP-[ H]Gal as described by Whiteheart et al. [30]. The labeled glycoproteins were then separated by a combination of non-equilibrium pH gel electrophoresis followed by SDS-PAGE as described by O Farrell [31]. The gels were impregnated with sodium salicylate, dried and the labeled glycoproteins were visualized by fluorography for the indicated times. Protein molecular weight markers are as indicated.

DNA electrophoresis results. A gel after DNA electrophoresis. Lane M is molecular weight markers. The numbers along the left-hand side are the sizes, in bp. at the marker molecule. 

FIGURE 7. Determination of the molecular weight of the purified PSPBP from Acanthocardia tuberculatum foot by gel electrophoresis on denaturing conditions. Molecular weight marker proteins were commercial myosine (205 kDa), P-galactosidase (116 kDa), phosphorylase B (97.4 kDa), albumin (66 kDa), ovalbumin (45 kDa) and carbonic anhydrase (29 kDa). The plot represents the relative mobilities of proteins vs. Log (Molecular Weight). 

Bacterial strains bearing plasmids of known sizes, to serve as molecular weight markers during electrophoresis, were obtained from Drs. P. Walsh and G. Jacoby. Lysates of these were prepared by the method described above. 

Fig. 1 Purification of amylase300. Panel A shows purification steps of amylase300. Panel B shows SDS-12% polyacrylamide gel electrophoresis of culture supernatant (lanel), purified amylase300 (lane 2) and molecular weight marker (lane MW).

Fig. 22.2 Summary of the extraction procedure for the SIPC. Bands for molecular weight markers on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) are to the left, and to the right, SIPC subunits. After Clare and Matsumura (2000) reproduced by permission of Taylor Francis Ltd. (www.informaworld.com)...

FIGURE 2. SDS-polyacrylamide gel electrophoresis of active fractions from purification of the D1 protease from pea. Proteins were detected by silver staining. M, molecular weight markers lane 1, crude extract from sonicated thylakoids lane 2, eluate from Fractogel TSK DEAE-650 column lane 3, eluate from TSK G2000 SWG column lane 4, eluate from TSK DEAE-5PW column. The putative protease, based on its apparent molecular weight from size exclusion chromatography, is arrowed. 

Fig. 4. SDS-polyacrylamide gel electrophoresis of poly(ADP-ribos)ylated DNA ligase II. Poly(ADP-ribos)ylation was carried out as described in Sect. 2 except that the NAD" concentration was changed as indicated, a Purified poly(ADP-ribose) polymerase b purified DNA ligase II c a negative control sample incubated without NAD d another negative control sample incubated with 13 jug/0.2 ml of purified poly(ADP-ribose) in place ofNAD e-h the samples incubated with 0.2, 0.5, 1, and 2 mM NAD", respectively / molecular weight markers. The position of poly(ADP-ribose) polymerase (P) and DNA ligase II (L) are indicated by arrows. The position of the latter enzyme was determined fluorographically by a parallel run of [ H]AMP-DNA ligase complex [12,13]...

Fig. 3. (left) SDS-PAGE of p PJNAD treated SR vesicles in the absence and presence of poly L-lysine. 0.88 mg/ml of SR was incubated at 25 C for 60 min with 20 [xM [32p]NAD (2.8 ci/mmol) and 0, 50 or 100 M,g/ml of poly L-lysine. Radiolabeling of the acid insoluble fraction (25 p.g protein) from each sample was analyzed by SDS-polyacrylamide gel electrophoresis (8). Tlie Coomassie brilliant blue-staining pattern (A) and an autoradiogram of the same gel (B) are shown. Molecular weight markers phosphorylase b (94 K), bovine serum albumin (67 K), ovalbumin (43 K), carbonic anhydrase (30 K) and soybean trypsin inhibitor (20.1 K). 

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