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| 선택 | Cat.No. | 제품명 | 가격(VAT별도) | 수량 |
|---|
제품특징
□ 특징
● Designed for cell-free DNA: newly formulated repair andligation reagents with optimized protocols
● High performance NGS libraries: high diversitywith broad and reproducible GC coverage
● Variable sample input: <1 ng to 30 ng of cell-freeDNA
● Fast and simple workflow: 3 steps in a single tube orwell in 2 hours with no purification or sample transfer steps
● Research applications: libraries from samples whereresults require high sensitivity including liquid biopsies, ctDNA, and targetedsequencing
● Automation-friendly: Beckman® FXP Workstation
□ 제품설명
ThruPLEX®Plasma-seq Kit은 혁신적인 ThruPLEX chemistry를바탕으로 plasma로부터 추출한 cell-free DNA를이용한 NGS용 library제작에 최적화된 제품으로, 본 제품을 이용하면 다양성이 높고,GC representation 유지하는 고퀄리티의 library를제작할 수 있다. 본 제품으로 제작한 library는 CNV analysis, genome sequencing application 뿐 아니라 AgilentSureSelect®, Roche Nimblegen® SeqCap® EZ 등 주요 Target Enrichment platform과같이 사용할 수 있다.
□ 보관
Store at -20 °C.Guarantee for 9 months at -20°C in a constant temperature freezer.
□ 내용
Component name | Cap Color | 12S Kit 12 Single Indexes | 48S Kit48 Single Indexes | 96D Kit96 Dual Indexes |
Code |
| RB4490 | RB4491 | RB4492 |
용량 |
| 12회 | 48회 | 96회 |
Preparation Buffer | Red | 1 Tube | 1 Tube | 2 Tubes |
Template Preparation Enzyme | Red | 1 Tube | 1 Tube | 2 Tubes |
Library Synthesis Buffer | Yellow | 1 Tube | 1 Tube | 2 Tubes |
Library Synthesis Enzyme | Yellow | 1 Tube | 1 Tube | 2 Tubes |
Library Amplification Buffer | Green | 1 Tube | 1 Tube | 2 Tubes |
Library Amplification Enzyme | Green | 1 Tube | 1 Tube | 2 Tubes |
Nuclease-Free Water | Clear | 1 Tube | 1 Tube | 1 Tube |
Indexing Reagents | Blue | 12 Tubes | 1 Single Index Plate (48S) | 1 Dual Index Plate (96D) |
※ ThruPLEX® Plasma-seq Kit single Index 서열 * ThruPLEX Plasma-seq 12S Kit (RB4490) * ThruPLEX Plasma-seq 48S Kit (RB4491)
[그림1] ThruPlex® Plasma-Seq KitSingle-tube WorkflowStartingwith 1 to 30 ng of cell-free DNA, ThruPLEX® Plasma-seq Kit creates indexed libraries in 3 simple steps: end repair, adapterligation, and high-fidelity library amplification. No purification or sampletransfer steps are required. The streamlined workflow is performed in 2 hoursin a single tube or well, preventing sample loss and enhancing positive sampleidentification.
[그림2] ThruPlex® Plasma-Seq Kit원리 ThruPLEX® Plasma-seq Kit technology is a3-step reaction that is optimized for cell-free DNA. Cell-free DNA (1 ng to 30ng) is first repaired in a highly efficient process. Background is reducedusing double-stranded adapters with no single-stranded tails. Blunt-endligation occurs with high-efficiency. Blocked 5’ ends reduce adapter-adapter ligation. Background is furtherreduced by destroying unused adapters after ligation. A high-fidelityamplification completes the reaction to generate indexed Illumina libraries.
[그림3] Highest Diversity, FewestUnmapped Reads from Cell-free DNA ThruPLEX® Plasma-seq Kit generated quality libraries with high diversity and a lownumber of duplicates and unmapped reads.Cell-free DNA was extracted from 3plasma samples, and libraries were prepared at the amounts indicated asmeasured by Qubit®.The amount of mononucleosomal DNA in each sampleas measured by the Bioanalyzer® corresponded to 0.09 ng, 0.62 ng, and 15.44 ng.Pooled libraries were sequenced on an Illumina NextSeq® 500 as apaired-end run with 17M to 25M reads per library, Duplicationrates were calculatedafter down-sampling the data to 17 M reads per library.
[그림4] OutstandingTarget EnrichmentPerformance ThruPLEX® Plasma-seq Kit libraries were captured at high efficiency and generateddata with deep coverage of the kinomefor mutation detection. Libraries wereprepared from 3 plasma samples at input amounts of 5 ng, 6.5 ng, and 10 ng in triplicate, and targeted sequencing was carried out on an Illumina MiSeq® using samples enriched with theClearSeq HumanDNA Kinome Panel for SureSelectXT2. On average, 5M reads weregenerated per library. Selected bases were successfullycaptured bases thatwere in or within 250 bp of the baits.
[그림5] Reproducible, Unbiased GCCoverage ThruPLEX® Plasma-seq Kit provided the mostreproducible and unbiased GC coverage across the human genome. ThruPLEX® librariesshowed minimalvariability across 9 individual plasma samples tested. Libraries were preparedfrom cell-free DNA isolatedfrom 1 ml of plasma samples and sequenced on anIllumina NextSeq® 500. Four separate plasma samples were used to constructthe NEBNext® Ultra libraries.
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3-硝基邻苯二甲酸可作为鲁米诺的合成原料。3-硝基邻苯二甲酸与肼在高沸点溶剂（如二甘醇）中发生缩合反应，失去一分子水，生成3-硝基邻苯二甲酰肼。然后以保险粉还原3-硝基邻苯二甲酰肼中的硝基，得到3-氨基邻苯二甲酰肼，即是鲁米诺。
我们常说的鲁米诺试剂是鲁米诺与过氧化氢（双氧水的主要成分）的混合物，主要用于现代刑侦的的血液检测。鲁米诺与氢氧化物反应时生成了一个双负离子（Dianion），它可被过氧化氢分解出的氧气氧化，产物为一个有机过氧化物。该过氧化物很不稳定，立即分解出氮气（鲁米诺被有机氧化剂如二甲基亚砜氧化后不是生成氮气，而是生成含氮有机物），生成激发态的3-氨基邻苯二甲酸。激发态至基态转化中，释放的能量以光子的形式存在，波长位于可见光的蓝光部分。鲁米诺只有用氧化剂处理过才会发光。通常使用双氧水和一种氢氧化物碱的混合水溶液作为激发剂。在铁化合物催化下，双氧水分解为氧气和水：2H2O2→O2+2H2O实验室中常以铁氰化钾作为催化剂铁的来源，而法医学上的催化剂则恰好是血红蛋白中的铁。很多生物系统中的酶也可催化过氧化氢的分解反应。鲁米诺试剂使用识别血液的试剂，血迹即使被擦拭，血液中的血红素还是会残留下来，当鲁米诺试剂喷在血红素上，会与活性氧产生氧化作用，释放出蓝紫色荧光。被成为鲁米诺反应。是鉴定血液用的有机物质。
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有荧光为何还需要压片这么久呢？不知各位用的时候有无这种情况！
配胶、上样、跑胶、半干转无特殊，半干转后，就用TBST浸润一下后放入封闭液（试过5%牛奶、5%BSA）2小时，后取出直接抗体孵育（目标蛋白抗体1：1000、1：1500、1：2000【建议浓度1：1000-3000】，二抗1：2000、1：3000、1：5000【建议浓度1：1000-5000】，内参beta-actin差不多，目标蛋白4度过夜，内参室温2小时），孵育完后，TBST洗三次，每次10min，后二抗孵育，完后同样TBST洗三次，每次10min，后进行ECL发光液浸润2min，暗室压片2min(主要问题是在暗室中可以看到整张PVDF膜都有荧光，郁闷），显影定影，结果在胶片PVDF膜位置很黑，虽然可以见到我的条带，但是无法使用，不知道究竟怎样解决？（实验室另一位同学不存这样的问题）
试过几次都是同样的问题，是封闭的不好？抗体稀释度不够？TBST洗得不好？ECL发光步骤问题？渴求园里战友指导，谢谢！
钡离子焰色反应为黄绿色
钙离子燃烧砖红色
铷离子焰色反应为紫色
铜离子焰色反应为绿色
钠离子焰色反应为黄色
钾离子焰色反应为浅紫色
锶离子焰色反应为洋红色
经常使用这些元素的硝酸盐作为焰火发色剂。
