{"product_id":"neb-m7634l","title":"New England Biolabs, M7634L, NEBNext UltraShear®","description":"Enzymatic fragmentation of DNA as part of the library prep workflow provides many advantages compared to mechanical shearing. However, specialized fragmentation reagents are required for enzymatic shearing in order to maintain methylation marks on samples for methylome analysis or for use with challenging samples such as FFPE DNA.  \u003cb\u003eRelated Categories\u003c\/b\u003e DNA Fragmentation \u0026amp; RNA Fragmentation,, FFPE DNA,, Next Generation Sequencing Library Preparation  \u003cb\u003eSpecification\u003c\/b\u003e \u003cb\u003eMaterials Required but not Supplied\u003c\/b\u003e 1X TE (10 mM Tris pH 8.0, 1 mM EDTA) 0.2 ml thin wall PCR tubes Magnetic rack\/stand (NEB #S1515S; Alpaqua® #A001322 or equivalent) PCR machine Vortex Microcentrifuge Bioanalyzer®, TapeStation® or other fragment analyzer and associated consumables 80% Ethanol For use with NEBNext UltraShear Protocol: SPRIselect™ Reagent Kit (Beckman Coulter, Inc. #B23317), AMPure® XP Beads (Beckman Coulter, Inc. #A63881) or Monarch®® PCR \u0026amp; DNA Cleanup Kit (NEB# T1030S\/L) are recommended for Section 1. For use with NEBNext Ultra II End Repair\/dA-Tailing Module Protocol: NEBNext Ultra II End Repair\/dA-Tailing Module (NEB #E7546S\/L) for Section 2. Recommended Material Not Included: NEBNext Ultra II Ligation Module (NEB #E7595) and NEBNext Multiplex Oligos (www.neb.com\/oligos). For use with NEBNext Enzymatic Methyl-seq Protocol: NEBNext Enzymatic Methyl-seq Kit (NEB #E7120S\/L) for Section 3. Formamide (Sigma #F9037-100 ml) or optional 0.1 N NaOH. Formamide is preferred. If using NaOH, please see NEBNext Enzymatic Methyl-seq Kit (NEB #E7120) FAQs Nuclease-free  \u003cb\u003eFAQ\u003c\/b\u003e Q: Is NEBNext UltraShear™ the same as NEBNext® Ultra™ II FS or NEBNext dsDNA Fragmentase®? A: No. These enzymatic fragmentation reagents do not contain the same enzymes. NEBNext UltraShear has been designed for challenging samples including FFPE DNA and for methylome analysis. NEBNext Ultra II FS is our recommendation for standard DNA library prep for Illumina® sequencing, and it has a more streamlined workflow. NEBNext dsDNA Fragmentase is our legacy product for enzyme-based DNA fragmentation. Q: Do you really need to vortex NEBNext UltraShear™? A: Yes, failure to vortex NEBNext UltraShear may produce variable results. Q: For EM-seq™ workflows, what are the recommendations for fragmenting already-fragmented DNA, low integrity DNA and\/or FFPE DNA with NEBNext UltraShear™? A: If working with already fragmented, low integrity and\/or FFPE DNA upstream of EM-seq library preparation (Section 5 of NEBNext UltraShear manual), for Step 5.1.6. we suggest fragmenting with NEBNext UltraShear for less time (5-15 minutes at 37˚C; optimizations may be needed). Additionally, we suggest performing 0.9X sample purification bead cleanups after Deamination of Cytosines (Step 5.9.2.) and after PCR Amplification (Step 5.11.3). Please note that we do not recommend a stringent sample purification bead cleanup (e.g., 0.65X) for already fragmented, low integrity and\/or FFPE DNA samples. Q: Following the fragmentation step of the NEBNext UltraShear™ protocol, can the reactions be stored at -20˚C? A: Yes, following NEBNext UltraShear fragmentation the reactions can be stored overnight at -20˚C. Q: Do you recommend NEBNext UltraShear™ for high-sensitivity, low error rate DNA library preparation? A: Use of NEBNext UltraShear for fragmentation has been reported to result in high sensitivity and low error rate compared to other enzymatic fragmentation methods and mechanical shearing (e.g. Covaris®), upstream of DNA library preparation. However, we do not have internal data for this application. Q: Can I fragment my gDNA sample with NEBNext UltraShear® in buffers other than 1X TE (10mM Tris pH 8.0, 1mM EDTA)? A: gDNA samples can be fragmented with NEBNext UltraShear® with common gDNA extraction kits’ elution buffers, e.g. Monarch® DNA Elution Buffer, Monarch® gDNA Elution Buffer, Monarch® gDNA Elution Buffer II, Buffer EB and Buffer AE, in place of 1X TE pH 8.0. However, the fragmentation for NEBNext UltraShear is slower with gDNA samples fragmented in these elution buffers. Fragmentation conditions would need to be optimized and fragmentation may need to be incubated for extended times at 37˚C or 45 ˚C compared to gDNA samples in 1X TE pH 8.0. Monarch® DNA Elution Buffer (NEB #T1016): 10 mM Tris, 0.1 mM EDTA pH 8.5 Monarch® gDNA Elution Buffer (NEB #T3016): 10 mM Tris, 0.1 mM EDTA pH 9.0 Monarch® gDNA Elution Buffer II (NEB #T3056) or Buffer AE: 10 mM Tris-HCl, 0.5 mM EDTA pH 9.0 Qiagen Buffer EB: 10 mM Tris-HCl pH 8.5 Q: If the fragmentation at 37˚C takes a long time (30 minutes or more), is there a way to speed up my NEBNext UltraShear fragmentation? A: If you are not achieving desired fragmentation profile after prolonged incubation at 37˚C, to speed up fragmentation, you can incubate the NEBNext UltraShear reaction at 45˚C instead of 37˚C. We do not recommend starting your optimization with 45˚C, since most samples will fragment at 37˚C. Using the higher temperature right away may over fragment the DNA. We recommend testing and optimization of these conditions on your specific sample type and application. ","brand":"New England Biolabs","offers":[{"title":"Default Title","offer_id":46835537969321,"sku":"M7634L","price":0.99,"currency_code":"USD","in_stock":true}],"url":"https:\/\/iright.com\/products\/neb-m7634l","provider":"Iright","version":"1.0","type":"link"}