polyacrylamide-agarose gel

Andre Hamel hamel at cc.umanitoba.ca
Thu Jun 2 17:25:38 EST 1994


I'm interested in any info on recipes, experiences, etc involving agarose-
acrylamide composite gels for nucleic acid electrophoresis (small
fragments, such as 100- 500 bp PCR products).

I recently posted that I'll search Uncover and Medline.

I received several requests, thus I'm posting references regarding
agarose- polyacrylamide gels.

Using Uncover (telnet database.carl.org) one key paper turned up (I've yet
to receive it by interlibrary loan, so if anybody reading this is familiar
with it please post experiences) ... Schiffer, R. 1992. "Clear as a gel".
Laboratory Practice. Feb.1 Vol 41, No 2, pg 9.

Using Medline I dug up following (excuse incomplete keywords ... a quirk
of our online system due to downloading).

Best luck all. ... any new info/experiences, etc, please keep us all
posted :^)

Andre

Here's Medline "mess";
SilverPlatter 3.1 __________________          MEDLINE EXPRESS (R) 1987________________________

MEDLINE EXPRESS (R) 1987________________________ usage is subject to the terms and conditions of the 
Subscription and License Agreement and the applicable Copyright and 
intellectual property protection as dictated by the appropriate laws of your 
country and/or by International Convention.

                                MEDLINE EXPRESS (R) 1989-1993           1 of 5  
                                                           Marked in Search: #6
TI: Electrophoresis_______________ of DNA in ultrathin planar-format linear polyacrylamide.
AU: MacDonell-MT; Roszak-DB
AD: Ransom Hill Bioscience, Ramona, CA 92065.
SO: Genet-Anal-Tech-Appl. 1993 Feb; 10(1): 10-5
ISSN: 1050-3862
PY: 1993
LA: ENGLISH
CP: UNITED-STATES
AB: Linear or un-cross-linked polyacrylamides have been employed successfully 
in the field of capillary electrophoresis_______________ for the separation of nucleic acids. 
Typical acrylamide__________ concentrations for those applications range from 3% to 14% 
(wt/vol), with consistencies ranging from virtually liquid to moderately 
viscous. Due to the absence of cross-links, and the relatively fluid nature of 
linear polyacrylamide at typically employed concentrations, its use in planar 
(slab) gel electrophoresis_______________ has been overlooked. We describe herein the 
application of ultrathin (100 microns) high-viscosity slabs of linear 
polyacrylamide to planar electrophoresis_______________ of nucleic acid fragments. The 
approach we describe is rapid and yields high-resolution separations of nucleic 
acid fragments in linear polyacrylamide supports. The mobilities of DNA 
fragments of various lengths in a range of concentrations of linear polymer are 
compared with those observed for conventional cross-linked gels. The reptative 
migration of larger DNA fragments in linear polymers is predictable from the 
models derived from work with cross-linked acrylamide__________ and agarose_______. The 
migration of smaller fragments, however, is not entirely predicted by the 
Ogston model. The relative mobilities observed for very small_____ DNA fragments are 
approximately half those predicted by the Ogston regimen.
MESH: Acrylamides-chemistry
MESH: *DNA-analysis; *Electrophoresis,-Polyacrylamide-Gel-methods___________________________________________
TG: Support,-U.S.-Gov't,-Non-P.H.S.
PT: JOURNAL-ARTICLE
RN: 0; 79-06-1; 9007-49-2
NM: Acrylamides; acrylamide__________; DNA
AN: 93319797
UD: 9310

                                MEDLINE EXPRESS (R) 1989-1993           2 of 5  
                                                           Marked in Search: #6
TI: DNA digest analysis with capillary electrophoresis_______________.
AU: Paulus-A; Husken-D
AD: Corporate Analytical Research, Ciba-Geigy Ltd., Basal, Switzerland.
SO: Electrophoresis_______________. 1993 Jan-Feb; 14(1-2): 27-35
ISSN: 0173-0835
PY: 1993
LA: ENGLISH
CP: GERMANY
AB: DNA digest analysis with polymer-filled capillaries in capillary 
electrophoresis_______________ is described. The samples analyzed consisted of commercially 
available standards including a 100 base pair ladder with repeating units up to 
2000 base pairs. Three DNA digests covered the most common small_____ fragment 
ranges: up to 600 base pairs, up to 1500 base pairs and from 100 to 2500 base 
pairs. All samples were separated by traditional slab gel electrophoresis_______________ at 
various agarose_______ concentrations and by automated capillary electrophoresis_______________. The 
capillary electrophoretic separations were achieved with noncross-linked 
polyacrylamide from 6% monomer solutions. The acrylamide__________ was polymerized inside 
the capillary, which was coated with a methacryloxysilane to insure binding of 
the polyacrylamide to the capillary wall. With 6% columns, excellent 
separations were observed up to 600 base pairs with base line resolution for 
fragments differing in less than 10 base pairs. Resolution power for fragments 
between 600 and 2200 decreased to about 300 base pairs. Compared to slab gels, 
capillary electrophoresis_______________ achieved better resolution in the low fragment range, 
whereas with the reported column composition, slab gels were superior above 600 
base pairs. Fast access to the analysis of a specific sample, automation, and a 
larger dynamic range for sample load are further benefits of a DNA digest 
analysis by capillary electrophoresis_______________.
MESH: Autoanalysis-; Bacteriophage-phi-X-174-genetics; Capillarity-; 
Deoxyribonucleases,-Type-II-Site-Specific-metabolism; DNA-metabolism; 
DNA,-Bacterial-isolation-and-purification; DNA,-Bacterial-metabolism; 
DNA,-Viral-isolation-and-purification; DNA,-Viral-metabolism; 
Electrophoresis,-Agar-Gel_________________________; Escherichia-coli-genetics; 
Repetitive-Sequences,-Nucleic-Acid
MESH: *DNA-isolation-and-purification; 
*Electrophoresis,-Polyacrylamide-Gel-methods___________________________________________
TG: Support,-Non-U.S.-Gov't
PT: JOURNAL-ARTICLE
RN: EC 3.1.21.-; EC 3.1.21.-; EC 3.1.21.-; EC 3.1.21.-; EC 3.1.21.4; 0; 0; 
9007-49-2
NM: endodeoxyribonuclease-BglI; endodeoxyribonuclease-HaeII; 
endodeoxyribonuclease-HinfI; endodeoxyribonuclease-HpaII; 
Deoxyribonucleases,-Type-II-Site-Specific; DNA,-Bacterial; DNA,-Viral; DNA
AN: 93215605
UD: 9307

                                MEDLINE EXPRESS (R) 1988                3 of 5  
                                                          Marked in Search: #13
TI: Discontinuous polyacrylamide______________ gradient agarose_______ gels resolve a wide range of 
restriction fragments and optimize the efficiency of nucleic acid transfer.
AU: Jones-CL; Simpson-NJ; Waryas-VL; Pappas-MG
AD: Covalent Technology Corporation, Ann Arbor, Michigan 48106.
SO: Anal-Biochem. 1988 Sep; 173(2): 285-8
ISSN: 0003-2697
PY: 1988
LA: ENGLISH
CP: UNITED-STATES
AB: A vertical electrophoresis_______________ procedure utilizing a discontinuous 
polyacrylamide______________ gradient agarose_______ gel was developed to resolve DNA___ fragments 
ranging in size from over 50 kb to less than 300 bp in length. The gel 
consisted of a polyacrylamide______________ plug at the base of the gel followed by a 
gradient of agarose_______ ranging from 0.3 to 0.9%. Restriction fragments migrated 
shorter distances than in a comparable polyacrylamide-0.3__________________% agarose_______ gel, and 
small_____ fragments were retained. Southern transfer of DNA___ fragments from the 
gradient gel onto nitrocellulose was more efficient than transfer of fragments 
using a nongradient gel.
MESH: Blotting,-Northern; Blotting,-Southern; Electrophoresis,-Agar-Gel_________________________; 
Electrophoresis,-Polyacrylamide-Gel___________________________________
MESH: *DNA-analysis____________; *Restriction-Mapping
TG: Support,-Non-U.S.-Gov't
PT: JOURNAL-ARTICLE
RN: 9007-49-2
NM: DNA___
AN: 89048303
UD: 8902

                                MEDLINE EXPRESS (R) 1988                4 of 5  
                                                          Marked in Search: #13
TI: Detection of base mutations in genomic DNA___ using denaturing gradient gel 
electrophoresis_______________ (DGGE) followed by transfer and hybridization with 
gene-specific probes.
AU: Borresen-AL; Hovig-E; Brogger-A
AD: Department of Genetics, Norwegian Radium Hospital, Oslo, Norway.
SO: Mutat-Res. 1988 Nov; 202(1): 77-83
ISSN: 0027-5107
PY: 1988
LA: ENGLISH
CP: NETHERLANDS
AB: It has been shown that minor differences, such as single-base-pair 
substitutions between otherwise identical DNA___ fragments can result in altered 
melting behavior detectable by denaturing gradient gel electrophoresis_______________ (DGGE). 
Sequence variations in only a small_____ DNA___ region within one locus can be detected 
using the previously described procedures. We have developed a method for the 
efficient Southern transfer of genomic DNA___ fragments from the denaturing 
gradient gels in order to be able to analyze larger regions in several loci for 
variation. The gels were made using polyacrylamide______________ containing 2% 
low-geling-temperature agarose_______ (LGT). The polyacrylamide______________ gel (PAG) was 
crosslinked with a reversible crosslinker, and after electrophoresis_______________ the 
crosslinks were cleaved, the structure of the gel being maintained by the 
agarose_______. After this treatment of the denaturing gels, more than 90% of the DNA___ 
fragments could be transferred to nylon membranes by alkaline transfer, while 
electroblotting transferred only 10% of the DNA___. Hybridization with 
gene-specific probes was then performed. We have used this technique to 
identify an RFLP in the COL1A2 gene in a human genomic DNA___ sample. The transfer 
technique described should make the use of DGGE more widely applicable since 
the genomic DNA___ fragments separated on one gel can be screened with several 
different probes, both cDNA and genomic probes.
MESH: Base-Sequence; Blotting,-Southern; DNA-Probes__________
MESH: *Collagen-genetics; *DNA-Mutational-Analysis-methods_______________________________; 
*Electrophoresis,-Polyacrylamide-Gel-methods___________________________________________; *Nucleic-Acid-Denaturation; 
*Polymorphism-Genetics; *Restriction-Fragment-Length-Polymorphisms
TG: Human-; Support,-Non-U.S.-Gov't
PT: JOURNAL-ARTICLE
RN: 0; 9007-34-5
NM: DNA-Probes__________; Collagen
AN: 89039989
UD: 8902

                                MEDLINE EXPRESS (R) 1987                5 of 5  
                                                          Marked in Search: #13
TI: Improved resolution of DNA___ fragments in polysaccharide-supplemented agarose_______ 
gels.
AU: Perlman-D; Chikarmane-H; Halvorson-HO
SO: Anal-Biochem. 1987 May 15; 163(1): 247-54
ISSN: 0003-2697
PY: 1987
LA: ENGLISH
CP: UNITED-STATES
AB: The electrophoretic separation of nucleic acids, including small_____ DNA___ 
fragments in the range 50-1000 bp, is presently carried out in polyacrylamide______________ 
gels or in gels containing high concentrations of agarose_______. We have developed an 
alternative gel matrix composition which is inexpensive, nontoxic, easy to 
prepare, and highly transparent to visible and uv light. The composition 
combines a soluble nonionic polysaccharide such as hydroxyethylcellulose, 
methylcellulose, or galactomannan with a minimum but sufficient concentration 
of agarose_______ to form a gel which immobilizes the "liquid phase sieve." These 
mixtures do not replace polyacrylamide______________ for resolving fragments smaller than 
approximately 75 nucleotides. However, the new gels show DNA___ fragment 
resolution (band separation versus distance traveled) and optical clarity 
superior to those of conventional agarose_______.
MESH: Carbohydrates-; Cellulose-analogs-and-derivatives; 
Electrophoresis,-Agar-Gel-methods_________________________________; Mannans-; Methylcellulose-; Polymers-
MESH: *DNA-analysis____________
TG: Support,-Non-U.S.-Gov't; Support,-U.S.-Gov't,-P.H.S.
PT: JOURNAL-ARTICLE
CN: GM18904
RN: 0; 0; 11078-30-1; 9004-34-6; 9004-62-0; 9004-67-5; 9007-49-2
NM: Mannans; Polymers; galactomannan; Cellulose; hydroxyethylcellulose; 
Methylcellulose; DNA___
AN: 87296706
UD: 8711
SilverPlatter 3.1 __________________          MEDLINE EXPRESS (R) 1987________________________

MEDLINE EXPRESS (R) 1987________________________ usage is subject to the terms and conditions of the 
Subscription and License Agreement and the applicable Copyright and 
intellectual property protection as dictated by the appropriate laws of your 
country and/or by International Convention.

                                MEDLINE EXPRESS (R) 1/94-5/94           1 of 5  
                                                          Marked in Search: #20
TI: Comparison of properties of agarose_______ for electrophoresis_______________ of DNA___.
AU: Upcroft-P; Upcroft-JA
AD: Queensland Institute of Medical Research, Bancroft Center, Herston 
Australia.
SO: J-Chromatogr. 1993 Aug 25; 618(1-2): 79-93
ISSN: 0021-9673
PY: 1993
LA: ENGLISH
CP: NETHERLANDS
AB: Agarose_______ as a medium for separation of DNA___ was first introduced in 1962 and 
since the early 1970s agarose_______ submarine gel electrophoresis_______________ has been synonymous 
with separations of DNA___ molecules larger than 1 kilobase pair (kb). The large 
pore size, low electroendosmosis and strength of the matrix have advantages 
over other media such as polyacrylamide______________ for many applications. The variety of 
grades of agarose_______, developed by chemical manipulation of the substitutions on 
the agarose_______ polymer, provides a range of matrices for separation of DNA___ 
molecules from a few base pairs (bp) to over 5 megabase pairs (Mb) in length. 
The introduction of low-melting-temperature agarose_______ has revolutionised the 
extraction and manipulation of chromosome-sized molecules. On the other hand, 
the demand for analysis of very small_____ quantities of DNA___ will most likely lead 
to the increasing importance of capillary electrophoresis_______________. Many theories have 
been propounded to explain the electrophoretic migration of DNA___ in agarose_______. The 
most popular of these has been reptation theory but none can account for all of 
the reported anomalies in migration. However, anomalous migration has been 
exploited to study DNA___ structure, topology and catenation. An example of the 
use of two-dimensional electrophoresis_______________ to demonstrate the complexity of DNA___ 
migration through agarose_______ is given. Generally, for molecules smaller than 50 
kb, electrophoretic separation is a function of length. By alternately 
electrophoresing DNA___ in two different directions, molecules as large as 5.7 Mb 
have been effectively separated, although with such large molecules DNA___ 
structure as well as size may determine migration. In the case of separations 
of chromosomes from the intestinal protozoan, Giardia duodenalis, for example, 
a discrepancy of 1 Mb in the size of one chromosome, with an apparent size of 
0.7-2.0 Mb, depended on the boundary conditions of separation. Major challenges 
for the molecular biologist are separation of larger chromosomal sized 
molecules, greater number of samples and smaller formats. Towards this 
challenge computer-aided technology is a key component in the control of 
electrophoresis_______________ parameters and analysis.
MESH: DNA,-Protozoan-analysis_______________________; Giardia-
MESH: *DNA-analysis____________; *Electrophoresis,-Agar-Gel-methods_________________________________; *Sepharose-standards
TG: Animal; Comparative-Study; Human
PT: JOURNAL-ARTICLE; REVIEW; REVIEW,-TUTORIAL
RN: 0; 9007-49-2; 9012-36-6
NM: DNA,-Protozoan______________; DNA___; Sepharose
AN: 94043642
UD: 9402

                                MEDLINE EXPRESS (R) 1989-1993           2 of 5  
                                                          Marked in Search: #20
TI: Advances in DNA___ electrophoresis_______________ in polymer solutions.
AU: Tietz-D; Aldroubi-A; Pulyaeva-H; Guszczynski-T; Garner-MM; Chrambach-A
AD: Laboratory of Theoretical and Physical Biology, National Institute of Child 
Health and Human Development, National Institutes of Health, Bethesda, MD 
20892-0001.
SO: Electrophoresis_______________. 1992 Sep-Oct; 13(9-10): 614-6
ISSN: 0173-0835
PY: 1992
LA: ENGLISH
CP: GERMANY
AB: DNA___ electrophoresis_______________ in gels and solutions of agarose_______ and polyacrylamide______________ was 
objectively evaluated with regard to separation efficiency at optimal polymer 
concentrations. In application to DNA___ fragments, polyacrylamide______________ gels were 
superior for separating fragments of less than 7800 bp, and agarose_______ gels are 
the best choice for larger fragments. Agarose_______ solutions are nearly as good as 
polyacrylamide______________ gels for small_____ DNA___ (< 300 bp). Agarose_______ solutions have a higher 
efficiency than polyacrylamide______________ solutions for DNA___ of less than 1200 bp. 
Separation efficiency sharply decreases with increasing length of DNA___. 
Retardation in polyacrylamide______________ solutions was found to depend on polymer length 
in a biphasic fashion. The choice of resolving polymer concentrations depends 
on the progressive stretching of DNA___ in proportion to polymer concentration. 
The rate of that stretching appears higher in polyacrylmide solution than in 
gels or in liquid or gelled agarose_______. Application of polymer solutions to 
capillary electrophoresis_______________ raises further problems concerning agarose_______ plugs, DNA___ 
interactions with the polymers, operation at low field strength and long 
durations as well as detection sensitivity.
MESH: Acrylic-Resins; DNA-chemistry_____________; Evaluation-Studies; Particle-Size; 
Sepharose-; Solutions-
MESH: *DNA-isolation-and-purification______________________________; *Electrophoresis,-Agar-Gel-methods_________________________________; 
*Electrophoresis,-Polyacrylamide-Gel-methods___________________________________________
TG: Comparative-Study
PT: JOURNAL-ARTICLE
RN: 0; 0; 0; 9007-49-2; 9012-36-6
NM: polyacrylamide-gels___________________; Acrylic-Resins; Solutions; DNA___; Sepharose
AN: 93092911
UD: 9303

                                MEDLINE EXPRESS (R) 1988                3 of 5  
                                                          Marked in Search: #20
TI: Discontinuous polyacrylamide______________ gradient agarose_______ gels resolve a wide range of 
restriction fragments and optimize the efficiency of nucleic_______ acid transfer.
AU: Jones-CL; Simpson-NJ; Waryas-VL; Pappas-MG
AD: Covalent Technology Corporation, Ann Arbor, Michigan 48106.
SO: Anal-Biochem. 1988 Sep; 173(2): 285-8
ISSN: 0003-2697
PY: 1988
LA: ENGLISH
CP: UNITED-STATES
AB: A vertical electrophoresis_______________ procedure utilizing a discontinuous 
polyacrylamide______________ gradient agarose_______ gel was developed to resolve DNA___ fragments 
ranging in size from over 50 kb to less than 300 bp in length. The gel 
consisted of a polyacrylamide______________ plug at the base of the gel followed by a 
gradient of agarose_______ ranging from 0.3 to 0.9%. Restriction fragments migrated 
shorter distances than in a comparable polyacrylamide-0.3__________________% agarose_______ gel, and 
small_____ fragments were retained. Southern transfer of DNA___ fragments from the 
gradient gel onto nitrocellulose was more efficient than transfer of fragments 
using a nongradient gel.
MESH: Blotting,-Northern; Blotting,-Southern; Electrophoresis,-Agar-Gel_________________________; 
Electrophoresis,-Polyacrylamide-Gel___________________________________
MESH: *DNA-analysis____________; *Restriction-Mapping
TG: Support,-Non-U.S.-Gov't
PT: JOURNAL-ARTICLE
RN: 9007-49-2
NM: DNA___
AN: 89048303
UD: 8902

                                MEDLINE EXPRESS (R) 1988                4 of 5  
                                                          Marked in Search: #20
TI: Detection of base mutations in genomic DNA___ using denaturing gradient gel 
electrophoresis_______________ (DGGE) followed by transfer and hybridization with 
gene-specific probes.
AU: Borresen-AL; Hovig-E; Brogger-A
AD: Department of Genetics, Norwegian Radium Hospital, Oslo, Norway.
SO: Mutat-Res. 1988 Nov; 202(1): 77-83
ISSN: 0027-5107
PY: 1988
LA: ENGLISH
CP: NETHERLANDS
AB: It has been shown that minor differences, such as single-base-pair 
substitutions between otherwise identical DNA___ fragments can result in altered 
melting behavior detectable by denaturing gradient gel electrophoresis_______________ (DGGE). 
Sequence variations in only a small_____ DNA___ region within one locus can be detected 
using the previously described procedures. We have developed a method for the 
efficient Southern transfer of genomic DNA___ fragments from the denaturing 
gradient gels in order to be able to analyze larger regions in several loci for 
variation. The gels were made using polyacrylamide______________ containing 2% 
low-geling-temperature agarose_______ (LGT). The polyacrylamide______________ gel (PAG) was 
crosslinked with a reversible crosslinker, and after electrophoresis_______________ the 
crosslinks were cleaved, the structure of the gel being maintained by the 
agarose_______. After this treatment of the denaturing gels, more than 90% of the DNA___ 
fragments could be transferred to nylon membranes by alkaline transfer, while 
electroblotting transferred only 10% of the DNA___. Hybridization with 
gene-specific probes was then performed. We have used this technique to 
identify an RFLP in the COL1A2 gene in a human genomic DNA___ sample. The transfer 
technique described should make the use of DGGE more widely applicable since 
the genomic DNA___ fragments separated on one gel can be screened with several 
different probes, both cDNA and genomic probes.
MESH: Base-Sequence; Blotting,-Southern; DNA-Probes__________
MESH: *Collagen-genetics; *DNA-Mutational-Analysis-methods_______________________________; 
*Electrophoresis,-Polyacrylamide-Gel-methods___________________________________________; *Nucleic-Acid-Denaturation_________________________; 
*Polymorphism-Genetics; *Restriction-Fragment-Length-Polymorphisms
TG: Human-; Support,-Non-U.S.-Gov't
PT: JOURNAL-ARTICLE
RN: 0; 9007-34-5
NM: DNA-Probes__________; Collagen
AN: 89039989
UD: 8902

                                MEDLINE EXPRESS (R) 1987                5 of 5  
                                                          Marked in Search: #20
TI: Improved resolution of DNA___ fragments in polysaccharide-supplemented agarose_______ 
gels.
AU: Perlman-D; Chikarmane-H; Halvorson-HO
SO: Anal-Biochem. 1987 May 15; 163(1): 247-54
ISSN: 0003-2697
PY: 1987
LA: ENGLISH
CP: UNITED-STATES
AB: The electrophoretic separation of nucleic_______ acids, including small_____ DNA___ 
fragments in the range 50-1000 bp, is presently carried out in polyacrylamide______________ 
gels or in gels containing high concentrations of agarose_______. We have developed an 
alternative gel matrix composition which is inexpensive, nontoxic, easy to 
prepare, and highly transparent to visible and uv light. The composition 
combines a soluble nonionic polysaccharide such as hydroxyethylcellulose, 
methylcellulose, or galactomannan with a minimum but sufficient concentration 
of agarose_______ to form a gel which immobilizes the "liquid phase sieve." These 
mixtures do not replace polyacrylamide______________ for resolving fragments smaller than 
approximately 75 nucleotides. However, the new gels show DNA___ fragment 
resolution (band separation versus distance traveled) and optical clarity 
superior to those of conventional agarose_______.
MESH: Carbohydrates-; Cellulose-analogs-and-derivatives; 
Electrophoresis,-Agar-Gel-methods_________________________________; Mannans-; Methylcellulose-; Polymers-
MESH: *DNA-analysis____________
TG: Support,-Non-U.S.-Gov't; Support,-U.S.-Gov't,-P.H.S.
PT: JOURNAL-ARTICLE
CN: GM18904
RN: 0; 0; 11078-30-1; 9004-34-6; 9004-62-0; 9004-67-5; 9007-49-2
NM: Mannans; Polymers; galactomannan; Cellulose; hydroxyethylcellulose; 
Methylcellulose; DNA___
AN: 87296706
UD: 8711




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