A simple apparatus for vertical flat-sheet polyacrylamide gel electrophoresis

Reid, M; Bieleski, R. L.

doi:10.1016/0003-2697(68)90278-9

Cited by 377 publications

(83 citation statements)

References 12 publications

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“…Acrylamide electrophoresis was carried out on all samples of each of the three systems ; on disc gels (Davis, 1964), with application of 5 µ uterine exúdate and 2-5 µ serum; gradient gels (Gradipore: Margolis & Kenrick, 1968) with 2-5 µ exúdate and serum; and slab gels (Reid & Bieleski, 1968) (Cowan & Daniel, 1972) and the wallaby (Renfree, 1973a).…”

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confidence: 99%

Uterine Proteins in the Marsupial, Didelphis Marsupialis Virginiana, During Gestation

Renfree¹

1975

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The No r t h American opossum, Didelphis marsupialis virginiana, is a polyovular, polyoestrous marsupial. The gestation period of 13 days is the same length as the luteal phase of the cycle (Hartman, 1923(Hartman, , 1925 (Hartman, 1923;Renfree, 1974). After ovulation, the two uteri increase greatly in size and turgidity, due to an accumulation of fluid in the hypertrophied mucosa and the development of a rich supply of capillaries and venules (Harman, 1923 (Renfree, 1974) were included for comparison.Opossums were collected and housed as already described (Renfree, 1974

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confidence: 99%

Uterine Proteins in the Marsupial, Didelphis Marsupialis Virginiana, During Gestation

Renfree¹

1975

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“…RNA was analyzed in slab gel systems (100 X 145 X 1.5 mm) (2,5,19) in 3.2 and 12% polyacrylamide gels. To separate large RNA molecules, 3.2% polyacrylamide gels were prepared in 0.04 M Trisacetate buffer (pH 7.6).…”

Section: Methodsmentioning

confidence: 99%

Colicin E3: A Unique Endoribonuclease

Meyhack

Apirion

1973

Proc. Natl. Acad. Sci. U.S.A.

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The question of whether or not a cellular ribonuclease is involved in the cleavage of 16S ribosomal RNA by colicin E3 was investigated. For this purpose ribosomes from strains devoid of some ribonucleases or ribosomes in which ribonucleases had been inactivated by heat or removed by extensive washings were used for the colicin reaction. Since the 16S RNA of all these different ribosomes, and even of the most extensively washed ribosomes, was cleaved by colicin E3, it is suggested that cellular ribonucleases are not involved in colicin E3 action.Thus, colicin E3 seems to be a unique endoribonuclease.Colicin E3 (1) cleaves 16S ribosomal RNA from Escherichia coli into two unequal fragments (2-4). The smaller fragment is about 50 nucleotides long and contains the 3' end of the molecule (2, 3). The cleavage reaction can take place in vivo after infection of sensitive E. coli cells with colicin (2, 3), or in vitro by incubation of ribosomes with colicin (4, 5). The reaction requires the presence of both ribosomal subunits (6, 7). Purified 16S ribosomal RNA is not cleaved by colicin E3 (2, 4). Moreover, colicin E3 has not been reported to possess any other ribonuclease activity.Under the reported conditions for in vitro colicin action on ribosomes (4,(6)(7)(8) there are a number of known, and perhaps several uhknown, ribonucleases that are still attached to the ribosomes. Therefore, it is possible that an adventitious ribonuclease is involved in colicin action. We shall consider here the following possibilities: (a) Colicin itself is the ribonuclease and has a unique substrate specificity; (b) Colicin activates a pre-existing ribonuclease to function as a very specific endoribonuclease; (c) Colicin, in conjunction with a ribosomal protein(s) becomes a ribonuclease, which is an elaboration of the first possibility. The experiments reported here suggest that either possibility a or c is the most likely explanation of colicin action. MATERIALS AND METHODSRibosomes from Escherichia coli strains D10 (8), MRE 600 (9), and N7060 (10) were used for in vitro colicin assays.These strains are all devoid of ribonuclease I (8-10). Strain N7060 is further characterized by a thermolabile ribonuclease II and a modified polynucleotide phosphorylase (10). Highly purified colicin E3 was kindly provided by Dr. D. R. Helinski (this preparation did not have any measurable ribonuclease activity). Colicin E3 activity was assayed according to Herschman and Helinski (11), by use of the colicin-sensitive strain D10. Kodak x-ray film (NS2T, 12.5 X 17.5 cm) was used for autoradiography. Carrier-free H332PO4 (50 mCi/ml) was purchased from New England Nuclear Corp. 156Isolation of Ribosomes from Colicin-Sensitive Cells. Cells of strain D10 were grown at 370 in 500 ml of mimimal medium (per liter: 14.5 g Tris, -100 mg MgSO4, 500 mg NaCl, 150 mg KCl, 500 mg NH4C1, adjusted to pH 7.4 with HCl) containing 0.2% glucose, 25 mg of L-methionine, and 16 mg of KH2PO4. All subsequent steps in the preparation of ribosomes were done at 4°. Cells were gro...

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“…All solutions were then heated to 50'C for 2.5 minutes, then immediately chilled on ice, to reduce the possiblity of RNA aggregation (6,7). Flat gel electrophoresis was performed using the apparatus and general procedure of Rdid and Bieleski (8). Acrylamide and methylenebisacrylamide recrystallization, gel solutions and electrophoresis buffer were :prepared according to Loening (9).…”

Section: Materials and Methods (2)mentioning

confidence: 99%

ISOLATION OF HIGH MOLECULAR WEIGHT RNAs FROM CULTURED NEURAL TUMOR CELLS.

Edwards¹,

Vellis²

1971

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A simple apparatus for vertical flat-sheet polyacrylamide gel electrophoresis

Cited by 377 publications

References 12 publications

Uterine Proteins in the Marsupial, Didelphis Marsupialis Virginiana, During Gestation

Uterine Proteins in the Marsupial, Didelphis Marsupialis Virginiana, During Gestation

Colicin E3: A Unique Endoribonuclease

ISOLATION OF HIGH MOLECULAR WEIGHT RNAs FROM CULTURED NEURAL TUMOR CELLS.

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