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商品详细NEB/Monarch® Genomic DNA Purification Kit/50 preps/T3010S
NEB/Monarch® Genomic DNA Purification Kit/50 preps/T3010S
NEB/Monarch® Genomic DNA Purification Kit/50 preps/T3010S
商品编号: T3010S
市场价: ¥0.00
美元价: 0.00
产地: 美国(厂家直采)
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产品分类: 其他
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联系Q Q: 3392242852
电话号码: 4000-520-616
电子邮箱: info@ebiomall.com
商品介绍

The Monarch Genomic DNA Purification Kit is a comprehensive solution for cell lysis, RNA removal, and purification of intact genomicDNA (gDNA) from a wide variety of biological samples, including cultured cells, blood, and mammalian tissues. Additionally, bacteriaand yeast can be processed with extra steps to enhance lysis in these tough-to-lyse samples. Protocols are also included to enablepurification from clinically-relevant samples such as saliva and cheek swabs as well as rapid cleanup of previously extracted gDNA.Purified gDNA has high quality metrics, including A260/A280 > 1.8 and A260/A230 > 2.0, high DIN scores and minimal residual RNA. Thepurified gDNA is suitable for downstream applications such as end-point PCR, qPCR and library prep for NGS. It typically hasa peak size of > 50kb, making this kit an excellent choice upstream of long-read sequencing platforms.

Workflow for the Monarch Genomic DNA Purification Kit

Specifications:

Input:

Cultured mammalian cells: up to 5 x 106cells

Mammalian whole blood: 100 µl

Tissue: up to 25 mg, depending on tissue type

Bacteria: up to 2 x 109

Yeast: up to 5 x 107

Saliva: up to 500 µl

Buccal swabs

Genomic DNA requiring cleanup

Binding Capacity: 30 µg genomic DNA
Yield: Varies depending on sample type
Genomic DNA Size: Peak size > 50 kb for most sample types; may be lower for saliva and buccal swabs
RNA Content< 1% (with included RNase A treatment)
Elution Volume≥35 µl, but 100 µl is recommended
Purity: A260/280 ≥ 1.8A260/230 ≥ 2.0

Validated Sample Types:

  • Mouse Tail
  • Mouse Ear
  • Mouse Liver
  • Rat Liver
  • Mouse Kidney
  • Mouse Spleen
  • Mouse Heart
  • Mouse Lung
  • Mouse Brain
  • Rat Brain
  • Mouse Muscle
  • Rat Muscle
  • Deer Muscle
  • Human Blood
  • Mouse Blood
  • Rabbit Blood
  • Pig Blood
  • Guinea Pig Blood
  • Cow Blood
  • Horse Blood
  • Dog Blood
  • Chicken Blood
  • HeLa Cells
  • HEK293 Cells
  • NIH3T3 Cells
  • E. coli
  • Rhodobacter sp.
  • B. cereus
  • T. kodakarensis
  • S. cerevisiae
  • Saliva
  • Buccal swab
Figure 1: The Monarch Genomic DNA Purification Kit efficiently purifies high-quality, high molecular weight gDNA from avariety of sample types.
100 ng of genomic DNA from each sample was loaded on a 0.75% agarose gel. gDNA was isolated following the standard protocols forblood, cultured cells and tissue, and the supplemental protocols for buccal swabs, saliva, Gram– and Gram+ bacteria. Starting materialused: 1 x 106 HeLa cells, 100 μl human blood, 10 μl bird blood, 10 mg frozen tissue powder, 1 buccal swab, 500 μl saliva and ~1 x 109 bacterial cells. Lambda DNA-Hind III digest (NEB #N3012) was used as a marker in the last lane (M). Purified gDNA samples wereanalyzed using a Genomic DNA ScreenTape® on an Agilent Technologies® 4200 TapeStation®. Samples typically yield peak sizes 50–70 kband DINs of ~9. The cell fractions processed in the buccal swab and saliva preps contain dead cells, as expected, causing a smear likepattern with typical low molecular weight apoptotic bands.
Figure 2: The Monarch Genomic DNA Purification Kit provides excellent yields for tissues that are problematic for other commercial kits.
Duplicate 10 mg samples of RNAlater®-stabilized rat tissue were cut to small pieces and subsequently lysed and purified according to the protocols provided for each kit. Optional RNase A steps were included. Elution was carried out with 100 µl elution buffer provided in the respective kits. Yields displayed are averages of the duplicate samples, and represent the genomic DNA yield after correcting for the RNA content as determined by LC-MS. Results indicate that the Monarch Genomic DNA Purification Kit provides excellent yields for a wide range of tissues.
Figure 3: Genomic DNA purified with the Monarch Genomic DNA Purification Kit is high quality and suitable for sensitive applications like long range PCR and qPCR.
A. Amplification reactions were set up with primer pairs specific for 6, 8, 10, 12, 16, 20 kb amplicons from human DNA. LongAmp® Hot Start Taq 2X Master Mix (NEB #M0533) was used and 25 ng template DNA was added to each sample. PCR reactions were carried out on an Applied Biosystems 2720 Thermal Cycler. Monarch-purified genomic DNA isolated from HeLa cells and human blood were compared to commercially available reference DNA from the human cell line NA19240 F11. 10 of 20 µl was loaded on a 1.5% agarose gel, using the 1 kb DNA Ladder (NEB #N3232) as a marker. Results indicated DNA was of high-integrity and suitable for long range PCR. B. Monarch-purified genomic DNA from human whole blood, HeLa cells and mouse tail was diluted to produce a five log range of input template concentrations. The results were generated using primers targeting gHEME (human whole blood) and gREL (HeLa, mouse tail) for qPCR assays with the Luna Universal qPCR Master Mix (NEB #M3003) and cycled on a Bio-Rad® CFX Touch qPCR thermal cycler. Results indicated that DNA is highly pure and free from inhibitors, optimal for qPCR.
Figure 4: The Monarch Genomic DNA Purification Kit produces excellent input material for NGS library preparation with NEBNext® kits for Illumina®.
A. Duplicate libraries were made from 100 ng HeLa cell gDNA purified with Monarch (orange) or Qiagen DNeasy Mini Kit (blue) using the NEBNext Ultra II FS DNA Library Prep Kit for Illumina (NEB #E7805). Libraries were sequenced on an Illumina MiSeq. Reads were mapped using Bowtie 2.2.4 and GC coverage was calculated using Picard’s CollectGCBiasMetrics (v1.117). Expected normalized coverage of 1.0 is indicated by the horizontal grey line, the number of 100 bp regions at each %GC is indicated by the vertical grey bars, and the colored lines represent the normalized coverage for each library. Monarch GC coverage matched Qiagen DNeasy results. B. Yields of libraries produced by enzymatic shearing are equivalent when using Monarch-purified gDNA. Library yields of the samples described above were assessed on an Agilent Technologies® 2100 BioAnalyzer using a High Sensitivity DNA Kit.
Figure 5: DNA isolation from higher volume and amounts of blood using RBC Lysis based protocol for Monarch gDNA Purification Kit
DNA extracted using RBC lysis method achieves similar high yields and concentrations as direct methods, while allowing for extraction from higher volumes and amounts of blood. Genomic DNA samples were purified in triplicate from 3 days old fresh pig blood. For the Direct Lysis samples, the DNA from 100 µl pig blood was purified using the standard protocol for fresh mammalian bloodfrom the NEB #T3010kit. For the RBC Lysis samples, the indicated amount of pig blood was used in the protocol for up to 500 µl bloodusing the NEB #T3010kit and RBC Lysis Buffer (NEB #T3051).
Figure 6: The Monarch Genomic DNA Purification Kit generates high quality DNA for nanopore sequencing.
HeLa cell genomic DNA was extracted using either the Monarch Genomic DNA Purification Kit or the Qiagen DNeasy Blood & Tissue Kit. One microgram of purified DNA was used to prepare Oxford Nanopore Technology (ONT) sequencing libraries following the ONT 1D Ligation Sequencing Kit (SQK-LSK109) protocol without DNA fragmentation. Libraries were loaded on a GridION (Flow cell R9.4.1) and the data was collected for 48 hrs. Libraries produced using the Monarch gDNA exceeded the Qiagen libraries on common sequencing metrics including: A. total sequencing data collected, B. read quality, and C. read length. Data was generated using NanoComp (Bioinformatics, Volume 34, Issue 15, 1 August 2018, Pages 2666–2669).
This product is related to the following categories:
Genomic DNA Extraction & Purification,
Nucleic Acid Purification Products
This product can be used in the following applications:
Nucleic Acid Purification
品牌介绍
New England Biolabs(NEB)公司       NEB公司——三十多年的卓越品质NEB公司成立于二十世纪七十年代中期,拥有众多经验丰富的科学家,是生产生命科学试剂的领导者。目前,NEB为基因组研究提供最齐全的重组酶和天然酶,并且公司业务范围已延伸至蛋白质组学和药品开发领域。回顾三十余年来的历程,NEB公司作为先驱公司之一,为促进生物科技工业的发展做出了巨大的贡献。NEB美国总部乔迁新址后拥有最尖端的设备,有一座现代化的发酵中心及设备齐全的实验室,这些实验室主要用于产品生产、质量监控、产品开发和基础科研之用。作为首批以商业规模生产限制性内切酶的公司之一,NEB一直专注于内切酶的研究,并保持业内领先水平。NEB公司一贯坚持以科学为本的原则,公司生产的试剂因其高质量、高性价比享誉世界。重组酶NEB公司对酶的生产与其基础科研不能分开,通过对限制/修饰系统的克隆和过量表达方面的研究,使我们能够大大降低成本,改善产品质量。NEB已经成功克隆了180多种内切酶,其中大多是完全克隆,少数是部分克隆。目前,NEB可供应240多种内切酶,其中180多种可以重组酶形式提供,同时还有大量的应用途广泛的重组聚合酶和重组修饰酶。质量与客户服务NEB公司凭借着严格的质检程序、深入的基础科研以及不断开发的研发项目,承诺为全球科技人员提供高纯度的科研产品。直接与NEB总部联系或与NEB的国际网点联系,即可体验到NEB的个性化客户服务。公司内负责生产以及负责进行质量监控的科学家(产品负责人)也就是技术支持人员。他们为客户解答有关限制核酸内切酶、甲基化酶、以及其他DNA修饰酶、蛋白质修饰酶方面的问题。同时,公司的有机合成部门可提供linkers、引物、adaptors、探针以及寡核苷酸合成等方面的信息。研究人员也可为客户提供技术支持服务,这些研究人员在DNA测序、甲基化、克隆、过量表达、发酵、蛋白质纯化以及蛋白质分析方面都有很深造诣。基础科研NEB在分子生物学和寄生虫学方面的基础研究由公司内部的资深科学家负责。这些科学家在他们各自的领域都卓有成绩,经常在业内著名杂志上发表文献,指导博士后工作、为大学生提供暑期实习机会,而且经常受邀去当地学校进行演讲。NEB鼓励公司与外界合作,每周学术探讨会为其他科学家们提供了交流和展示的平台。环保政策NEB公司在资助科研项目时最基本原则之一就是:一切工作都应符合保护生态环境。公司产品生产、分析以及运输过程既要满足产品的稳定性,也要符合我们的环保政策。20年来,NEB公司最引人注目的一项环保政策是:回收运输泡沫盒,虽然这项工作首创于美国,但是,目前加拿大、德国以及英国的分公司也同样实施了。公司还在内部开展多方位的回收工作,经常用再生纸张印刷市场宣传资料。这些小细节不会影响产品质量,但是对环境的保护却是显而易见。公司职责除环保政策外,通过基金会,NEB还竭力为改善当地社区及全人类尽微薄之力。NEB基金会创立于1982年,属私人性质,其宗旨是支持发展中国家的环保、教育、健康及艺术事业。此外,NEB捐款委员会也经常向当地社区进行捐款活动。
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