SELECTION GUIDE – Nucleic Acid Purification

Function Product Name Page Major Subsequent Applications Capacity/Scale

Total RNA
Isolation of total RNA QuickPrep Total RNA Extraction 208 RT-PCR; Northerns; Up to 1 g tissue or
from cells or tissue Kit (27-9271-01) differential display 1 u 108 cells
Isolation of total RNA from RNA Extraction Kit 209 Northerns; mRNA purification; Up to 4 g tissue/kit
cells or tissue (27-9270-01) in vitro translation
Isolation of total RNA from Cesium Trifluoroacetate 209 Northerns; mRNA purification; 10 g tissue/100 ml
cells or tissue (17-0847-02) in vitro translation
mRNA
Purification of mRNA from QuickPrep Micro mRNA 210 RT-PCR; Northerns; cDNA 0.1 g tissue or 1 u 107 cells
cells or tissue Purification Kit (27-9255-01) synthesis; in vitro translation

Purification of mRNA from QuickPrep mRNA Purification Kit 210 Northerns; cDNA synthesis; 0.5 g tissue or 5 u 107 cells
cells or tissue (27-9254-01) in vitro translation; RT-PCR
Purification of mRNA from mRNA Purification Kit 211 cDNA synthesis; Northerns; 25 mg to 1 g cells or tissue
cells or tissue (27-9258-01, -02) in vitro translation; RT-PCR
Purification of mRNA from Oligo (dT)-Cellulose Type 7 211 cDNA synthesis; Northerns; Up to 4 mg poly(A)/1 g
cells or tissue (27-5543-01, -02, -03) in vitro translation; RT-PCR of cellulose
Newly Synthesized Oligonucleotides – featuring DNA-Grade Sephadex G-25 Resin
Desalting of oligonucleotides MicroSpin G-25 Columns 213 PCR; sequencing; labelling 100-150 µl of
x 10 bases (27-5325-01) deprotection solution

Desalting of oligonucleotides NAP Columns (17-0852, 17-0853, 214 PCR; sequencing; labelling Concentration
x 10 bases 17-0854; all -01, -02) w 1 mg/ml; 0.5-2.5 ml
Desalting of oligonucleotides Sephadex G-25, DNA Grade 213 PCR; labelling; sequencing Varies
x 10 bases (17-0572-01, -02)
Purification of oligonucleotides RESOURCE Q, RESOURCE 15RPC 495, 553 Sequencing; labelling; PCR Varies
(standard, modified, Columns (various code numbers)
phosphorothioate)
Labelled Oligonucleotides
Removal of unincorporated MicroSpin G-25 Columns 213 Hybridization 10-50 µl
label from end-labelled (27-5325-01)
oligonucleotides x 10 bases
DyeDeoxy Labelled DNA
Removal of excess DyeDeoxy AutoSeq G-50 (27-5340-01, -02, -03) 217 Automated sequencing on 12-25 µl
terminators from cycle ultra-salt-sensitive DNA
sequencing reactions sequencers (e. g. ABI 377 model)
Removal of excess DyeDeoxy AutoSeq96 (27-5340-10, -96) 218 High-throughput 10–20 µl/sample
terminators from cycle automated sequencing 96 samples/plate
sequencing reactions (e. g. MegaBACE 1000, ABI 3700)
Removal of excess DyeDeoxy MicroSpin G-50 (27-5350-01, -02) 219 Automated sequencing 12-25 µl
terminators from cycle
sequencing reactions
PCR Products and Restriction Fragments
Removal of excess primers MicroSpin S-400 HR Columns 215 PCR; cloning; sequencing 25-50 µl prior to
(w 32 bases) from PCR products (27-5140-01) subsequent PCR or
at least 200 bp in length cloning; 51-100 µl
prior to sequencing
Removal of excess primers MicroSpin S-300 HR Columns 215 Sequencing 25-50 µl
(w 20 bases) from PCR products (27-5130-01)
at least 100 bp in length
Buffer exchange MicroSpin G-25 Columns 213 Restriction 10-100 µl
(27-5325-01)

Purification and concentration GFX PCR DNA and Gel Band 216 Sequencing; restriction; cloning 100 µl reaction; 300 mg
of PCR products/DNA (100 bp- Purification Kit (27-9602-01) agarose per microfuge tube
10 kbp) from solution or
from gel bands
Purification of PCR products GFX 96 PCR Purification Kit 216 Labelling; automated and manual 20-100 µl
from solution (25-6902-01,-02) sequencing, restriction cloning
Purification of PCR products Sephaglas BandPrep Kit 217 Sequencing; restriction; cloning 0.75 g agarose per
from agarose; purification (27-9285-01) microfuge tube
of other DNAs from solution
Separation of PCR fragments Mono Q PC 1.6/5 Precision 493 Various Varies
or restriction fragments Column (17-0671-01)
Separation of PCR fragments Mono Q HR 5/5 Column 493 Various Varies
or restriction fragments (17-0546-01)
Typical Yield Total Time Format Advantages Notes
Total RNA (continued)
Mouse liver: 1 hour Microfuge (small scale) Speed; no hazardous Not for use with bacteria or whole blood
4.6 µg/mg tissue or standard centrifuge organic solvents
3 mg total RNA/g Overnight Ultracentrifuge Good for RNase-rich Ideal for low-copy-number messages
calf liver sources
Varies Overnight Ultracentrifuge Strong chaotrope; Can be used to separate RNA and DNA
inhibits ribonucleases
mRNA (continued)
Up to 6 µg/100 mg 15 minutes/round; Spin (batch); microfuge Speed; functionally pure For isolation of mRNA directly
tissue 1 hour for 24 samples message; high throughout from cells and tissue; not ideal
with MicroPlex 24 when used with MicroPlex 24 for low-copy-number messages
Vacuum Vacuum
up to 25 µg v 1 hour Spin-column; tabletop Speed; individually packed;
(90% purity) centrifuge functionally pure message
20 µg/600 µg total 30-45 minutes/ Spin-column; tabletop Individually packed
RNA (1 round) round centrifuge reagents
Varies Varies Spin; gravity; batch Economy, scalable

Newly Synthesized Oligonucleotides – featuring DNA-Grade Sephadex G-25 Resin (continued)

85-90% recovery 4 minutes/2 columns; Spin-columns; microfuge; Speed; prepacked; high
20 minutes/48 columns tabletop centrifuge throughput when used
with MicroPlex 24 with MicroPlex 24
90% recovery v 15 minutes Gravity Accommodates large
volumes (0.5-2.5 ml)
90% recovery Varies Spin or gravity Economy

Varies v 30 minutes High performance liquid Speed and purity; scalable For use with HPLC or with FPLC System
chromatography

Labelled Oligonucleotides (continued)

85-90% recovery 4 minutes/2 columns; Spin-column; microfuge Speed; high throughput Prepacked and pre-equilibrated
20 minutes/48 columns tabletop centrifuge when used with MicroPlex 24
with MicroPlex 24
DyeDeoxy Labelled DNA (continued)
80% recovery 4 minutes/2 columns; Spin-column; microfuge; Speed; high throughput Prepacked and pre-equilibrated
20 minutes/48 columns tabletop centrifuge when used with MicroPlex 24
with MicroPlex 24
65% recovery 15 minutes/96 samples 96-well microplate;tabletop centrifuge Speed, high throughput Prepacked and pre-equilibrated
with multichannel
pipettor
80% recovery 4 minutes/2 columns; Spin-column; microfuge; Speed; high throughput Prepacked and pre-equilibrated
20 minutes/48 columns tabletop centrifuge when used with MicroPlex 24
with MicroPlex 24
PCR Products and Restriction Fragments (continued)
50-70% recovery 4 minutes/2 columns; Spin-column; microfuge; Speed; high throughput Prepacked and pre-equilibrated
20 minutes/48 columns tabletop centrifuge when used with MicroPlex 24
with MicroPlex 24

60-80% recovery 4 minutes/2 columns; Spin-column; microfuge; Speed; high throughput Prepacked and pre-equilibrated
20 minutes/48 columns tabletop centrifuge when used with MicroPlex 24
with MicroPlex 24
90% recovery 4 minutes/2 columns; Spin-column; microfuge; Speed; high throughput Prepacked and pre-equilibrated
20 minutes/48 columns tabletop centrifuge when used with MicroPlex 24
with MicroPlex 24
90% from solution; 5 minutes from GFX spin-column; microfuge; Speed; purity; high throughput No glass slurry; elution in low-salt
60-80% from agarose solution; 15 minutes vacuum; tabletop centrifuge when used with MicroPlex 24 buffer; for agarose gel slices
from agarose Vacuum buffered in TBE or TAE

60-90% recovery 20min/96 samples 96-well plate; vacuum, centrifuge Speed; purity; high throughput No ethanol precipitation
no organic extractions
50% from agarose v 30 minutes for Spin (batch); microfuge Speed; scalable For purification of PCR products from
agarose plug agarose

Varies 10-30 minutes High performance Speed; small scale For use with SMART System
liquid chromatography
Varies v 30 minutes High performance Speed For use with HPLC or with FPLC System
liquid chromatography
SELECTION GUIDE (continued)
Function Product Name Page Major Subsequent Applications Capacity/Scale
Labelled dsDNA – featuring DNA-Grade Sephadex G-50 Resin
Removal of unincorporated ProbeQuant G-50 Micro Columns 218 Hybridization 25-50 µl for oligo-
label from labelled DNA (27-5335-01) labelled DNA; 25-100 µl
x 20 bases for end-labelled DNA
Removal of unincorporated MicroSpin G-50 Columns 219 Hybridization 25-50 µl for oligo-
label from labelled DNA (27-5330-01, -02) labelled DNA
x 20 bases
Removal of unincorporated MicroSpin S-200 HR Columns 219 Hybridization 51-100 µl
label from labelled DNA
x 50 bases
Removal of unincorporated NICK Columns 219 Hybridization Concentration
label from labelled DNA (17-0855-01, -02) w 1 mg/ml; w 100 µl
x 20 bases
Removal of unincorporated Sephadex G-50, DNA Grade 219 Hybridization Varies
label from labelled DNA (Grade F, 17-0573-01, -02)
x 20 bases
Plasmid DNA
Isolation of plasmid DNA GFX Micro Plasmid Prep Kit 220 Sequencing; restriction; labelling 1-3 ml cultures
from cultures (27-9601-01, -02)

Isolation of plasmid or cosmid FlexiPrep Kit (27-9281-01) 221 Sequencing; restriction; labelling 1.5-500 ml cultures
DNA from cultures

Isolation of plasmid DNA Sephacryl S-1000 (17-0476-01) 220 Sequencing; restriction; labelling Varies
from cultures
Buffer exchange of sequencing MicroSpin G-25 Columns 213 Sequencing 10-50 µl
templates (RNase-treated) (27-5325-01)

Purification of plasmid DNA/ MicroSpin S-300 HR Columns 215 Sequencing 25-50 µl

buffer exchange of sequencing
templates (not RNase-treated)
cDNA
Size selection of double-
stranded cDNA products
Genomic DNA
Isolation of genomic DNA from
whole blood, cultured cells,
lymphocytes, buccal cells,
yeast and bacteria.
Isolation of genomic DNA from
whole blood, bone marrow or
nucleated blood
Isolation of genomic DNA from
cultured cells, nucleated RBCs,
animal or plant tissue or
Gram-negative bacteria
Isolation of genomic DNA from
whole blood, buffy coat and
cultured cells
(27-5130-01)
SizeSep 400 Spun Columns
(27-5105-01)
GFX Genomic Blood DNA
Purification Kit (27-9603-01)
GenomicPrep Blood DNA
Isolation Kit (27-5236-01)
GenomicPrep Cells and Tissue
DNA Isolation Kit (27-5237-01)
Nucleon BACC (RPN8501/8502/8512)
221 Cloning cDNA made from up to
5 µg mRNA; 50-110 µl
222 PCR; sequencing; restriction; 10 µl-1 ml whole blood;
Southerns up to 4 u 106 cultured cells
223 PCR; sequencing; restriction; 300 µl-30 ml whole blood
Southerns
223 PCR; sequencing; restriction; 1 u 106 to 1 u 108
Southerns cultured cells or up to
0.5 g animal tissue
225 PCR; sequencing; restriction; 50 µl to 10 ml whole
Southerns blood; 1 u 106-1 u 107
cultured cells

Isolation of genomic DNA from Nucleon HT (RPN8509) 226 PCR; sequencing; restriction; 25 mg tissue sample;
animal tissue that is difficult to Southerns 30 µm paraffin section
homogenize (e.g. mouse tail,
bone) and tissue embedded
in paraffin
Isolation of genomic DNA from Nucleon PhytoPure (RPN8510/8511) 225 PCR; RAPDs; restriction Up to 0.1 g (small-scale);
plant tissue Southerns; AFLP up to 1.0 g (large-scale)
Typical Yield Total Time Format Advantages Notes
Labelled dsDNA – featuring DNA-Grade Sephadex G-50 Resin (continued)
80% recovery 4 minutes/2 columns; Spin-column; microfuge; Speed; high throughput Prepacked and pre-equilibrated;
20 minutes/48 columns tabletop centrifuge when used with MicroPlex 24 altermative to TCA precipitations;
with MicroPlex 24 sample elutes in 150 mM NaCI
80% recovery 4 minutes/2 columns; Spin-column; microfuge; Speed; high throughput Prepacked and pre-equilibrated;
20 minutes/48 columns tabletop centrifuge when used with MicroPlex 24 may also be used to desalt/buffer
with MicroPlex 24 exchange 10-100 µl samples
70-80% recovery 4 minutes/2 columns; Spin-column; microfuge; Speed; high throughput Prepacked and pre-equilibrated
20 minutes/48 columns tabletop centrifuge when used with MicroPlex 24
with MicroPlex 24
90% recovery v 15 minutes Gravity Speed; prepacked No centrifuge required

90% recovery Varies Spin or gravity Economy

Plasmid DNA (continued)

3-6 µg/ml culture 30 minutes for GFX spin-column; Speed; purity; high No glass slurry; elution in low-salt buffer
18 preps microfuge; vacuum throughput when used with
MicroPlex 24 Vacuum
y 2-4 µg/ml culture z20 minutes for a Spin (batch); microfuge; Economy; scalable Elution in low-salt buffer; can also
1.5 ml culture when tabletop centrifuge be used to isolate cosmid DNA
using streamlined
protocol
Varies Varies Gravity or pump Economy; isolation of Linear DNA v 20 kb is excluded
Form I plasmid from matrix
90% recovery 4 minutes/2 columns; Spin-column; microfuge; Speed; high throughput Prepacked and pre-equilibrated
20 minutes/48 columns tabletop centrifuge when used with MicroPlex 24
with MicroPlex 24
70-80% recovery 4 minutes/2 columns; Spin-column; microfuge; Speed; high throughput Prepacked and pre-equilibrated
20 minutes/48 columns tabletop centrifuge when used with MicroPlex 24
with MicroPlex 24
cDNA (continued)
Varies v 1 hour Spin-column; tabletop centrifuge; Prepacked Retains cDNAs v 400 bp; quality-control
Sepharose CL-4B resin tested for cDNA applications
Genomic DNA (continued)
Up to 15 µg 30 minutes for GFX spin-column; Speed; purity; high No glass slurry; elution in low-salt buffer
18 preps microfuge; vacuum; throughput when used with
tabletop centrifuge MicroPlex 24 Vacuum

35 µg/ml whole blood 2-3 hours Microfuge (small-scale); Purity; scalable

tabletop centrifuge
(large-scale)
35 µg/7 u 10 6 2-3 hours Microfuge (small-scale); Purity; scalable
nucleated cells tabletop centrifuge
(large-scale)

370-440 µg DNA/ 30 minutes Microfuge (small-scale); Purity; scalable; Novel protein binding resin; protocol
10 ml whole blood; benchtop centrifuge non-column means high modifications for buccal swabs,
8-12 µg DNA/106 (large-scale) mw DNA; no phenol mouthwashes, soft tissue, sperm,
cultured cells gram negative bacteria
70-200 µg DNA/cm 3 hours – o/n Microfuge Purity; non-column gives Novel protein binding resin
mouse tall; yields Proteinase K high yields
from sections digest followed by
not quantified 30 minutes extraction

Variable depending 1-1.5 hours Microfuge (small-scale) Purity; scalable; dedicated Novel polysaccharide binding resin
on species, age and benchtop centrifuge plant kit
part of plant used but (large-scale)
approximately 10-60 µg
per 0.1 g prep