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MagMAX Microbiome Ultra Nucleic Acid Isolation Kits
MagMAX Microbiome kit for faster and easier purification of high quality nucleic acids, with unique reagents specifically developed for microbiome research.
No matter your sample type and how difficult it is to lyse, the Applied Biosystems Microbiome kit can help you pull pure total nucleic acid for your research.
The benefits of the MagMAX Microbiome Ultra Nucleic Acid Isolation Kits include:
- Faster workflow—both DNA and RNA can be isolated in less than 1 hour
- Automation-ready—optimized to work with KingFisher instruments
- Efficient recovery—recover DNA/RNA from a wide range of micro-organisms without additional processing steps
- Compatible with human and environmental samples—Protocols for soil, stool, swabs, biofluids, and microbial cultures
 MagMAX Microbiome Ultra Nucleic Acid Isolation Kit |  Invitrogen PureLink Microbiome DNA Purification Kit | |
| Who should use? | Research labs that are developing their own tests for microbiome research and need both low- and high-throughput solutions for isolating total nucleic acid | Microbiome researchers processing 24 samples or less at a time |
| Sample types | Stool, swabs, transport media, culture media, urine, saliva, and soil | |
| Research needs |
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|
| KingFisher instrument compatibility | KingFisher Apex, Duo Prime, Flex, and Presto instruments | None (manual only with spin columns) |
| Elution volume | 50 μL to 200 μL | |
| Time per run | 96 stool samples processed in <60 minutes* | 96 stool samples processed in 120 minutes |
| Nucleic acid(s) | RNA and DNA | DNA only |
| Cat. No. | A42357, A42358 | A29790 |
| * Processing time for other sample types may vary. | ||
Multiple donors
Results using bead tubes
Figure 1. Quality of total microbial nucleic acid purified from feces (100 mg input) analyzed on a 1% agarose gel. 1 μg of total nucleic acid sample from each donor was loaded per lane in duplicate for six human donors. Clear bands of both genomic DNA and RNA can be seen on the ethidium bromide pre-stained agarose gel. The Invitrogen 1 Kb Plus DNA Ladder was utilized for size determination.
Note: Since this is not a denatured gel – RNA sizes shown are different from actual. Gel analysis was performed under native conditions, and thus RNA sizes appear somewhat different from the actual.
Figure 2. Total Nucleic acid yields (both DNA + RNA) isolated from fecal (100 mg input) samples collected from six human donors were measured on Nanodrop 2000.
Figure 3. Bioanalyzer trace of human fecal total RNA isolated using a bead tube version of MagMAX Microbiome ultra Total nucleic acid isolation kit. RNA 6000 Nano Kit with the Prokaryotic RNA assay was used to analyze the quality. Representative electropherogram indicate the position of 16S and 23S rRNA peaks. Unlabeled peaks correspond to additional ribosomal RNA.
Results using bead plate
Figure 4. Quality of total microbial nucleic acid purified from feces (100 mg input) analyzed on a 1% agarose gel. 1.5 μg of total nucleic acid sample from each donor was loaded per lane in duplicate for four human donors. Clear bands of both genomic DNA and RNA can be seen on the ethidium bromide pre-stained agarose gel. The Invitrogen 1 Kb Plus DNA Ladder was utilized for size determination.
Note: Since this is not a denatured gel – RNA sizes shown are different from actual. Gel analysis was performed under native conditions, and thus RNA sizes appear somewhat different from the actual.
Figure 5. Total Nucleic acid yields (both DNA and RNA) isolated from fecal (100 mg input) samples collected from four human donors were measured on Nanodrop 2000.
Figure 6. Bioanalyzer trace of human fecal total RNA isolated using a bead plate version of MagMAX Microbiome ultra Total nucleic acid isolation kit. RNA 6000 Nano Kit with the Prokaryotic RNA assay was used to analyze the quality. Representative electropherogram indicate the position of 16S and 23S rRNA peaks. Unlabeled peaks correspond to additional ribosomal RNA.
Expression analysis of DNA and RNA from bead plates
Figure 7. qPCR analysis of DNA purified from fecal samples with the MagMAX Microbiome Ultra Nucleic Acid Isolation Kit for four human donors.TaqMan Assays were utilized for one gram-positive (Firmicutes) and one gram-negative (Bacteroidetes) bacteria. 1:100 dilutions of total nucleic acid were used for Firmicutes and Bacteroidetes analysis in TaqMan Assays. TaqMan Fast Advanced Master Mix was used under fast cycling conditions.
Figure 8. qPCR analysis of RNA purified from fecal samples with the MagMAX Microbiome Ultra Nucleic Acid Isolation Kit for four human donors.TaqMan Assays were utilized for one gram-positive (Firmicutes) and one gram-negative (Bacteroidetes) bacteria. For RNA analysis Total nucleic acid was treated with DNase using DNA Free kit (Ambion) and reverse transcribed using Superscript Vilo Master Mix to make the cDNA. 1:100 dilutions of cDNA were used for Firmicutes and Bacteroidetes analysis in TaqMan Assays. TaqMan Fast Advanced Master Mix was used under fast cycling conditions.
Fecal swab using bead plate
Figure 9. Quality of total microbial nucleic acid purified from fecal swabs analyzed on a 1% agarose gel. 1 μg of total nucleic acid sample isolated from fecal swabs were loaded per lane. Two options were tested – adding direct swab in to the bead plate/tube well and swab in the transport media. Clear bands of both genomic DNA and RNA can be seen on the ethidium bromide pre-stained agarose gel (80 volts, 1 hr). The Invitrogen 1 Kb Plus DNA Ladder was utilized for size determination.
Note: Since this is not a denatured gel – RNA sizes shown are different from original. Gel analysis was performed under native conditions, and thus RNA sizes appear somewhat different from the actual.
Figure 10. Total Nucleic acid yields (both DNA and RNA) isolated from frozen fecal swabs and fecal swabs in transport media were measured on Nanodrop 2000.
Urine, saliva and VTM
Figure 11. qPCR analysis of DNA purified from Urine samples with the MagMAX Microbiome Ultra Nucleic Acid Isolation Kit for five human donors.TaqMan Assays were utilized for one gram-positive (Firmicutes) and one gram-negative (E. coli) bacteria. For RNA analysis-Total nucleic acid was treated with DNase using DNA Free kit (Ambion) and reverse transcribed using Superscript Vilo Master Mix to make the cDNA. TaqMan Fast Advanced Master Mix was used under fast cycling conditions.
Figure 12. qPCR analysis of DNA purified from Saliva samples with the MagMAX Microbiome Ultra Nucleic Acid Isolation Kit for five human donors.TaqMan Assays were utilized for one gram-positive (Firmicutes) and one gram-negative (E. coli) bacteria. For RNA analysis-Total nucleic acid was treated with DNase using DNA Free kit (Ambion) and reverse transcribed using Superscript Vilo Master Mix to make the cDNA. TaqMan Fast Advanced Master Mix was used under fast cycling conditions.
Figure 13. qPCR analysis of total nucleic acid isolated from Viral Transport Media (VTM) samples with the MagMAX Microbiome Ultra Nucleic Acid Isolation Kit. Zeptometrix spike-in controls were spiked-in to the VTM during the extraction process. TaqMan Assays were performed for several targets including (-)ssRNA virus, (+)ssRNA virus, dsDNA virus and a gram negative target that were included in the spike-in control mix. TaqMan Fast Virus 1-Step Master Mix was used under fast cycling conditions.
Fungal
Figure 14. qPCR analysis of DNA purified from fungal cultures with the MagMAX Microbiome Ultra Nucleic Acid Isolation Kit. qPCR was performed using a total fungal TaqMan assay. TaqMan Fast Advanced Master Mix was used under fast cycling conditions.
Figure 15. qPCR analysis of DNA purified from human donors with C. difficle infection using the MagMAX Microbiome Ultra Nucleic Acid Isolation Kit. Samples were obtained from Discovery biosciences and qPCR was performed using a C. difficile TaqMan assay. TaqMan Fast Advanced Master Mix was used under fast cycling conditions.
Mammals and other species
Figure 16. TaqMan assay for total nucleic acid isolated from fecal samples of mammals and reptiles with the MagMAX Microbiome Ultra Nucleic Acid Isolation Kit. TaqMan Assay was performed for a gram positive target (Firmicutes). TaqMan Fast Advanced Master Mix was used under fast cycling conditions.
Figure 17. qPCR analysis of total nucleic acid isolated from fecal samples of mammals, reptiles, bird, fish and fungal cultures with the MagMAX Microbiome Ultra Nucleic Acid Isolation Kit. TaqMan Assay was performed for mixed targets of both gram positive and gram negative (16S). TaqMan Fast Advanced Master Mix was used under fast cycling conditions.
Soil
Figure 18. qPCR analysis of total nucleic acid isolated from Texas soil samples with the MagMAX Microbiome Ultra Nucleic Acid Isolation Kit. TaqMan Assays were performed for a mixed gram positive and gram negative target (16S) and a Gram negative target (Bacteroidetes). Twenty fold dilution of soil Total nucleic acid preps were used for TaqMan assays. For RNA analysis-Total nucleic acid was treated with DNase using DNA Free kit (Ambion) and reverse transcribed using Superscript Vilo Master Mix to make the cDNA. TaqMan Fast Advanced Master Mix was used under fast cycling conditions.
Figure 19.Species level 16S profile for microbial DNA purified from stool of a human donor with Ulcerative Colitis. Total nucleic acid was purified from the stool of a patient with UC (in duplicates) and 16S Metagenomics kit and Ion plus fragment library kit were used to synthesize 16S libraries. The barcoded libraries were pooled and templated on Ion Chef Instrument followed by sequencing on the Ion S5 System. Automated analysis, annotation and taxonomic assignments were performed on Ion reporter software. A fecal sample DNA from a healthy donor is shown for species level comparison and abundance scale is shown above the profile. Red being the highly abundant species and blue being not detected/rare. R studio program is used to generate heat maps using log(2) values.
Click image to enlarge
Figure 20. Species level 16S profile for microbial DNA purified from stool of a human donor, who was on a nutritionist recommended diet for 16 weeks. Total nucleic acid was isolated from the stool of human donor who underwent (or was on) 16 weeks diet, in duplicate and 16S Metagenomics kit and Ion plus fragment library kit were used to synthesize 16S libraries. The barcoded libraries were pooled and templated on Ion Chef Instrument followed by sequencing on the Ion S5 System. Automated analysis, annotation and taxonomic assignments were performed on Ion reporter software. A fecal sample DNA from a no diet taking donor is shown for species level comparison and abundance scale is shown above the profile. Red being the highly abundant species and blue being not detected/rare. R studio program is used to generate heat maps using log(2) values.
Q: Which sample types can be processed with this kit?
A: The kit was developed for fast and efficient isolation of inhibitor-free microbial DNA/RNA from the human stool, swabs (rectal, skin, buccal) and body fluids (urine, saliva, VTM). It was also confirmed to perform well with rat stool and soil. Talking into account that stool and soil are the most challenging sample types in terms of homogenization and depletion of inhibitors, the kit should work well for most other sample types that researchers might be processing in the lab.
Q: Does this kit efficiently lyse the gram-positive bacteria and challenging fungal species?
A: The kit is utilizing a combination of chemical and mechanical (bead beating) lysis strategies, which allows to efficiently lyse even the most challenging members of the microbial community, and recover their DNA/RNA.
Q: Is the purified DNA/RNA completely inhibitor free?
A: The MagMax microbiome kit is based on a newly developed buffer system, allowing to eliminate vast majority of diverse inhibitors of enzymatic reactions, even from the most challenging samples such as stool and soil. In very rare instances, if some inhibitors still remain within the purified DNA/RNA, the sample can be diluted 10-20x prior to downstream PCR or other reactions.
Q: What are the bead beating options for this kit?
| Options for Bead beating | Settings |
|---|---|
| Omni Bead Ruptor 96 | 30 Hz - 2 min |
| Mini Bead Beater 96 | 2 min |
| Bead Bug | 4 min for 4 m/s |
| Plate Shaker (Vortex with plate adapter) | 5 min at 2000 rpm |
| Vortex with 24 tube adapter | 10 min at 2500 rpm |
Q: What are the options to obtain DNA only or RNA only using this kit?
For DNA only: Add 10 μL of RNase A (10 μg; AM2271) in each of the Wash 1 and Wash 2 plates.
For RNA only:DNA-free DNA Removal Kit (AM1906) can be used on purified total nucleic acid.
Multiple donors
Results using bead tubes
Figure 1. Quality of total microbial nucleic acid purified from feces (100 mg input) analyzed on a 1% agarose gel. 1 μg of total nucleic acid sample from each donor was loaded per lane in duplicate for six human donors. Clear bands of both genomic DNA and RNA can be seen on the ethidium bromide pre-stained agarose gel. The Invitrogen 1 Kb Plus DNA Ladder was utilized for size determination.
Note: Since this is not a denatured gel – RNA sizes shown are different from actual. Gel analysis was performed under native conditions, and thus RNA sizes appear somewhat different from the actual.
Figure 2. Total Nucleic acid yields (both DNA + RNA) isolated from fecal (100 mg input) samples collected from six human donors were measured on Nanodrop 2000.
Figure 3. Bioanalyzer trace of human fecal total RNA isolated using a bead tube version of MagMAX Microbiome ultra Total nucleic acid isolation kit. RNA 6000 Nano Kit with the Prokaryotic RNA assay was used to analyze the quality. Representative electropherogram indicate the position of 16S and 23S rRNA peaks. Unlabeled peaks correspond to additional ribosomal RNA.
Results using bead plate
Figure 4. Quality of total microbial nucleic acid purified from feces (100 mg input) analyzed on a 1% agarose gel. 1.5 μg of total nucleic acid sample from each donor was loaded per lane in duplicate for four human donors. Clear bands of both genomic DNA and RNA can be seen on the ethidium bromide pre-stained agarose gel. The Invitrogen 1 Kb Plus DNA Ladder was utilized for size determination.
Note: Since this is not a denatured gel – RNA sizes shown are different from actual. Gel analysis was performed under native conditions, and thus RNA sizes appear somewhat different from the actual.
Figure 5. Total Nucleic acid yields (both DNA and RNA) isolated from fecal (100 mg input) samples collected from four human donors were measured on Nanodrop 2000.
Figure 6. Bioanalyzer trace of human fecal total RNA isolated using a bead plate version of MagMAX Microbiome ultra Total nucleic acid isolation kit. RNA 6000 Nano Kit with the Prokaryotic RNA assay was used to analyze the quality. Representative electropherogram indicate the position of 16S and 23S rRNA peaks. Unlabeled peaks correspond to additional ribosomal RNA.
Expression analysis of DNA and RNA from bead plates
Figure 7. qPCR analysis of DNA purified from fecal samples with the MagMAX Microbiome Ultra Nucleic Acid Isolation Kit for four human donors.TaqMan Assays were utilized for one gram-positive (Firmicutes) and one gram-negative (Bacteroidetes) bacteria. 1:100 dilutions of total nucleic acid were used for Firmicutes and Bacteroidetes analysis in TaqMan Assays. TaqMan Fast Advanced Master Mix was used under fast cycling conditions.
Figure 8. qPCR analysis of RNA purified from fecal samples with the MagMAX Microbiome Ultra Nucleic Acid Isolation Kit for four human donors.TaqMan Assays were utilized for one gram-positive (Firmicutes) and one gram-negative (Bacteroidetes) bacteria. For RNA analysis Total nucleic acid was treated with DNase using DNA Free kit (Ambion) and reverse transcribed using Superscript Vilo Master Mix to make the cDNA. 1:100 dilutions of cDNA were used for Firmicutes and Bacteroidetes analysis in TaqMan Assays. TaqMan Fast Advanced Master Mix was used under fast cycling conditions.
Fecal swab using bead plate
Figure 9. Quality of total microbial nucleic acid purified from fecal swabs analyzed on a 1% agarose gel. 1 μg of total nucleic acid sample isolated from fecal swabs were loaded per lane. Two options were tested – adding direct swab in to the bead plate/tube well and swab in the transport media. Clear bands of both genomic DNA and RNA can be seen on the ethidium bromide pre-stained agarose gel (80 volts, 1 hr). The Invitrogen 1 Kb Plus DNA Ladder was utilized for size determination.
Note: Since this is not a denatured gel – RNA sizes shown are different from original. Gel analysis was performed under native conditions, and thus RNA sizes appear somewhat different from the actual.
Figure 10. Total Nucleic acid yields (both DNA and RNA) isolated from frozen fecal swabs and fecal swabs in transport media were measured on Nanodrop 2000.
Urine, saliva and VTM
Figure 11. qPCR analysis of DNA purified from Urine samples with the MagMAX Microbiome Ultra Nucleic Acid Isolation Kit for five human donors.TaqMan Assays were utilized for one gram-positive (Firmicutes) and one gram-negative (E. coli) bacteria. For RNA analysis-Total nucleic acid was treated with DNase using DNA Free kit (Ambion) and reverse transcribed using Superscript Vilo Master Mix to make the cDNA. TaqMan Fast Advanced Master Mix was used under fast cycling conditions.
Figure 12. qPCR analysis of DNA purified from Saliva samples with the MagMAX Microbiome Ultra Nucleic Acid Isolation Kit for five human donors.TaqMan Assays were utilized for one gram-positive (Firmicutes) and one gram-negative (E. coli) bacteria. For RNA analysis-Total nucleic acid was treated with DNase using DNA Free kit (Ambion) and reverse transcribed using Superscript Vilo Master Mix to make the cDNA. TaqMan Fast Advanced Master Mix was used under fast cycling conditions.
Figure 13. qPCR analysis of total nucleic acid isolated from Viral Transport Media (VTM) samples with the MagMAX Microbiome Ultra Nucleic Acid Isolation Kit. Zeptometrix spike-in controls were spiked-in to the VTM during the extraction process. TaqMan Assays were performed for several targets including (-)ssRNA virus, (+)ssRNA virus, dsDNA virus and a gram negative target that were included in the spike-in control mix. TaqMan Fast Virus 1-Step Master Mix was used under fast cycling conditions.
Fungal
Figure 14. qPCR analysis of DNA purified from fungal cultures with the MagMAX Microbiome Ultra Nucleic Acid Isolation Kit. qPCR was performed using a total fungal TaqMan assay. TaqMan Fast Advanced Master Mix was used under fast cycling conditions.
Figure 15. qPCR analysis of DNA purified from human donors with C. difficle infection using the MagMAX Microbiome Ultra Nucleic Acid Isolation Kit. Samples were obtained from Discovery biosciences and qPCR was performed using a C. difficile TaqMan assay. TaqMan Fast Advanced Master Mix was used under fast cycling conditions.
Mammals and other species
Figure 16. TaqMan assay for total nucleic acid isolated from fecal samples of mammals and reptiles with the MagMAX Microbiome Ultra Nucleic Acid Isolation Kit. TaqMan Assay was performed for a gram positive target (Firmicutes). TaqMan Fast Advanced Master Mix was used under fast cycling conditions.
Figure 17. qPCR analysis of total nucleic acid isolated from fecal samples of mammals, reptiles, bird, fish and fungal cultures with the MagMAX Microbiome Ultra Nucleic Acid Isolation Kit. TaqMan Assay was performed for mixed targets of both gram positive and gram negative (16S). TaqMan Fast Advanced Master Mix was used under fast cycling conditions.
Soil
Figure 18. qPCR analysis of total nucleic acid isolated from Texas soil samples with the MagMAX Microbiome Ultra Nucleic Acid Isolation Kit. TaqMan Assays were performed for a mixed gram positive and gram negative target (16S) and a Gram negative target (Bacteroidetes). Twenty fold dilution of soil Total nucleic acid preps were used for TaqMan assays. For RNA analysis-Total nucleic acid was treated with DNase using DNA Free kit (Ambion) and reverse transcribed using Superscript Vilo Master Mix to make the cDNA. TaqMan Fast Advanced Master Mix was used under fast cycling conditions.
Figure 19.Species level 16S profile for microbial DNA purified from stool of a human donor with Ulcerative Colitis. Total nucleic acid was purified from the stool of a patient with UC (in duplicates) and 16S Metagenomics kit and Ion plus fragment library kit were used to synthesize 16S libraries. The barcoded libraries were pooled and templated on Ion Chef Instrument followed by sequencing on the Ion S5 System. Automated analysis, annotation and taxonomic assignments were performed on Ion reporter software. A fecal sample DNA from a healthy donor is shown for species level comparison and abundance scale is shown above the profile. Red being the highly abundant species and blue being not detected/rare. R studio program is used to generate heat maps using log(2) values.
Click image to enlarge
Figure 20. Species level 16S profile for microbial DNA purified from stool of a human donor, who was on a nutritionist recommended diet for 16 weeks. Total nucleic acid was isolated from the stool of human donor who underwent (or was on) 16 weeks diet, in duplicate and 16S Metagenomics kit and Ion plus fragment library kit were used to synthesize 16S libraries. The barcoded libraries were pooled and templated on Ion Chef Instrument followed by sequencing on the Ion S5 System. Automated analysis, annotation and taxonomic assignments were performed on Ion reporter software. A fecal sample DNA from a no diet taking donor is shown for species level comparison and abundance scale is shown above the profile. Red being the highly abundant species and blue being not detected/rare. R studio program is used to generate heat maps using log(2) values.
Q: Which sample types can be processed with this kit?
A: The kit was developed for fast and efficient isolation of inhibitor-free microbial DNA/RNA from the human stool, swabs (rectal, skin, buccal) and body fluids (urine, saliva, VTM). It was also confirmed to perform well with rat stool and soil. Talking into account that stool and soil are the most challenging sample types in terms of homogenization and depletion of inhibitors, the kit should work well for most other sample types that researchers might be processing in the lab.
Q: Does this kit efficiently lyse the gram-positive bacteria and challenging fungal species?
A: The kit is utilizing a combination of chemical and mechanical (bead beating) lysis strategies, which allows to efficiently lyse even the most challenging members of the microbial community, and recover their DNA/RNA.
Q: Is the purified DNA/RNA completely inhibitor free?
A: The MagMax microbiome kit is based on a newly developed buffer system, allowing to eliminate vast majority of diverse inhibitors of enzymatic reactions, even from the most challenging samples such as stool and soil. In very rare instances, if some inhibitors still remain within the purified DNA/RNA, the sample can be diluted 10-20x prior to downstream PCR or other reactions.
Q: What are the bead beating options for this kit?
| Options for Bead beating | Settings |
|---|---|
| Omni Bead Ruptor 96 | 30 Hz - 2 min |
| Mini Bead Beater 96 | 2 min |
| Bead Bug | 4 min for 4 m/s |
| Plate Shaker (Vortex with plate adapter) | 5 min at 2000 rpm |
| Vortex with 24 tube adapter | 10 min at 2500 rpm |
Q: What are the options to obtain DNA only or RNA only using this kit?
For DNA only: Add 10 μL of RNase A (10 μg; AM2271) in each of the Wash 1 and Wash 2 plates.
For RNA only:DNA-free DNA Removal Kit (AM1906) can be used on purified total nucleic acid.
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