Genomic DNA from microoganisms - Macherey …

1 US:Tel.: +1 484 821 0984 Fax: +1 484 821 1272 E-mail: GmbH & Co. KG Neumann-Neander-Str. 6 8 52355 D ren / International:Tel.: +49 24 21 969-0 Fax: +49 24 21 969-199 E-mail: +41 62 388 55 00 Fax: +41 62 388 55 05 E-mail: +33 388 68 22 68 Fax: +33 388 51 76 88 E-mail: GmbH & Co. KG Neumann-Neander-Str. 6 8 52355 D ren / International:Tel.: +49 24 21 969-0 Fax: +49 24 21 969-199 E-Mail: +41 62 388 55 00 Fax: +41 62 388 55 05 E-Mail: +33 388 68 22 68 Fax: +33 388 51 76 88 E-Mail: ISO 9001 ZERTIFIZIERTMACHEREY-NAGEL GmbH & Co.

2 KG Neumann-Neander-Str. 6 8 52355 D ren / International:Tel.: +49 24 21 969-0 Fax: +49 24 21 969-199 E-Mail: +41 62 388 55 00 Fax: +41 62 388 55 05 E-Mail: +33 388 68 22 68 Fax: +33 388 51 76 88 E-Mail: ISO 9001 ZERTIFIZIERTG enomic DNA from microoganismsUser manualNucleoSpin Microbial DNAJuly 2018 / Rev. 03 Genomic DNA from microorganisms Protocol at a glance (Rev. 03) Macherey -NAGEL GmbH & Co. KG Neumann-Neander-Str. 6 8 52355 D ren GermanyTel.: +49 24 21 969-270 Fax: +49 24 21 969-199 Microbial DNA1 Prepare sample< 40 mg microbial pellet (wet weight)

3 100 L BE2 Lyse sampleTransfer sample in NucleoSpin Bead Tube Type B40 L Buffer MG10 L Liquid Proteinase KAgitate on a swing mill or similar device 4 12 min11,000 x g, 30 s3 Adjust binding conditions600 L Buffer MGVortex 3 s11,000 x g, 30 s4 Bind DNALoad 500 600 L sample on NucleoSpin Microbial DNA Column11,000 x g, 30 s5 Wash silica membrane1st2nd500 L BW 11,000 x g, 30 s500 L B5 11,000 x g, 30 s6 Dry silica membrane11,000 x g, 30 s7 Elute DNA100 L BERT, 1 min11,000 x g, 30 s 3 Macherey -NAGEL 07/2018, Rev.

4 03 Genomic DNA from microorganismsTable of contents1 Components Kit contents Reagents, consumables, and equipment to be supplied by user About this user manual 52 Product description The basic principle Kit specifications Handling, preparation, and storage of starting materials Lysis and disruption of sample material Disruption device 8 Type of Bead tube Time and frequency of disruption Elution procedures 103 Storage conditions and preparation of working solutions 104 Safety instructions 115 Protocols Protocol for gram positive and gram negative bacteria Protocol for yeast ( , Saccharomyces cerevisiae)

5 156 Appendix Troubleshooting Ordering information Product use restriction / warranty 19 Macherey -NAGEL 07/2018, Rev. 034 Genomic DNA from microorganisms1 Components Kit contentsNucleoSpin Microbial DNAREF10 preps preps preps Buffer MG10 mL38 mL5 x 38 mLWash Buffer BW6 mL30 mL150 mLWash Buffer B5 (Concentrate)*6 mL6 mL50 mLElution Buffer BE**13 mL30 mL125 mLLiquid Proteinase K120 L600 L2 x mLNucleoSpin Bead Tubes Type B1050250 NucleoSpin Microbial DNA Columns (light green rings)1050250 Collection Tubes (2 mL)20100500 User manual111* For preparation of working solutions and storage, see section 3.

6 ** Composition of Elution Buffer BE: 5 mM Tris/HCl, pH 07/2018, Rev. 03 Genomic DNA from Reagents, consumables, and equipment to be supplied by userReagents 96 100 % ethanolConsumables mL or 2 mL microcentrifuge tubes for microbial sample sedimentation Disposable tipsEquipment Manual pipettors Centrifuge for microcentrifuge tubes Vortex mixer Sample disruption device: swing mill or similar device ( , Mixer Mill MM200, MM300, MM400 (Retsch ); FastPrep System (MP-Biomedicals); Precellys (Bertin Technologies); MagNA Lyser (Roche); TissueLyser (QIAGEN); Bullet Blender (Next Advance).)

7 Mini-Beadbeater (Biospec Products); Speed Mill (Analytik Jena); Vortex Adapter for Vortex-Genie 2 X (MoBio)) Personal protection equipment (lab coat, gloves, goggles) About this user manualIt is strongly recommended for first time users to read the detailed protocol sections of the NucleoSpin Microbial DNA kit before using this product. Experienced users, however, may refer to the Protocol at a glance instead. The Protocol at a glance is designed to be used only as a supplemental tool for quick referencing while performing the purification technical literature is available online at contact Technical Service regarding information about any changes to the current user manual compared with previous 07/2018, Rev.

8 036 Genomic DNA from microorganisms2 Product The basic principleThe NucleoSpin Microbial DNA kit is designed for efficient isolation of Genomic DNA from microbial samples. DNA can be isolated from a wide variety of microorganisms such as gram negative, and gram positive bacteria as well as yeast, , Escherichia coli, Bacillus subtilis, Corynebacterium glutamicum, Saccharomyces cerevisiae. Preparation of the collected samples containing the microbes of interest should be in pellet data also indicate the usability of the kit for DNA isolation from fungal mycelia, , Aspergillus nidulans, from bacterial spore suspensions, , Geobacillus stearothermophilus, and from plant pollen, , honey bee pollen baskets.

9 For optimal DNA yield, bead tubes different from the ones included in the kit might be required for such applications (see section ).Microbial samples such as gram positive bacteria, yeast, and spores can be difficult to lyse due to their strong complex cell wall structures. The NucleoSpin Microbial DNA kit replaces enzymatic lysis by utilizing mechanical disruption of cell wall structures with the NucleoSpin Bead Tubes. The NucleoSpin Bead Tubes can be used in combination with many compatible disruptive devices (see section ). High DNA yields can be obtained with the NucleoSpin Bead Tubes from a large variety of sample types enabling the procedure to be convenient, fast, and easy.

10 Alternative bead types can be ordered separately for select sample types (see section for recommendations).7 Macherey -NAGEL 07/2018, Rev. 03 Genomic DNA from Kit specifications Kit specifications at a glanceParameterNucleoSpin Microbial DNAT echnologySilica membrane technologyFormatMini spin columnSample materialMicrobial cell culture pellets of gram positive and gram negative bacteria, yeastSample amountUp to approx. 40 mg wet weightTypical yieldVaries by sample and disruption device. 5 25 g DNA from approx. 30 mg wet weight microbial pellet can be obtainedA260 / volume100 200 LPreparation time35 min/6 prepsBinding capacity60 Handling, preparation, and storage of starting materialsCells should be harvested from fresh microbial cultures by sedimentation via centrifugation.