BIOLOVER

Decoding the Human Genome: The ENCODE Project and ChIP-seq Unveiled Decoding the Human Genome: The ENCODE Project and ChIP-seq Unveiled Have you ever wondered what makes us, well, us ? Beyond the familiar sequence of A, T, C, and G, our DNA holds a vast, intricate landscape of functional elements that dictate everything from our eye color to our susceptibility to diseases. For decades, scientists have grappled with the sheer complexity of the human genome, but thanks to groundbreaking initiatives like the ENCODE Project and powerful technologies such as ChIP-seq , we're finally beginning to unravel its deepest secrets. Ready to explore the hidden language of our genes? Let's dive in! The Grand Vision: Unveiling the ENCODE Project The ENCODE Project (Encyclopedia of DNA Elements) isn't just another scientific endeavor; it's a monumental undertaking with one ultimate goal: to b...

Mastering NGS Library Prep: Your Guide to Successful DNA Fragmentation & PCR Optimization Mastering NGS Library Prep Welcome to the cutting-edge world of bioinformatics! Have you ever wondered about the secrets behind successful Next-Generation Sequencing (NGS) experiments? NGS has revolutionized life science research, but let's be honest, its intricate process can be a bit daunting. And when it comes to NGS, library preparation is arguably the most crucial step. A tiny misstep here can jeopardize your entire experiment! That's why today, we're diving deep into two core components of NGS library preparation: DNA fragmentation and PCR cycle optimization . I'm here to share all my insights and help you navigate these critical stages, ensuring your NGS experiments are as successful as possible. Ready to unlock the full potential of your sequencing data? Let's get started! NGS Sequencing Success: It All Starts with DNA...

TEV Protease 처리와 Ni Spin Column 정제 가이드 MBP-tag 단백질 정제 를 할 때 많은 연구자들이 선택하는 시스템이 바로 pMAL-c6T 벡터 입니다. 이 시스템을 활용하면 MBP-tag와 표적 단백질을 효율적으로 분리할 수 있고, 특히 TEV protease를 이용한 절단 과정 과 이후 Ni Spin Column을 통한 분리 단계 는 높은 순도의 단백질을 확보하는 데 매우 중요합니다. 이번 포스트에서는 TEV 처리 후 정제 단계별로 버퍼 교환이 필요한지 여부 , 단백질 손실의 원인 , 그리고 실험 중 확인 방법(SDS-PAGE) 까지 꼼꼼히 안내드릴게요. TEV protease 처리 시 버퍼 교환은 필요할까? 많은 분들이 궁금해하시는 것 중 하나는, amylose resin에서 elution한 단백질 용액에 TEV protease를 처리하기 전 버퍼 교환이 필요한지 입니다. 정답은 NO! Amylose 컬럼의 elution buffer는 TEV protease의 반응 버퍼와 매우 유사 하기 때문에, 10X TEV buffer를 따로 첨가하지 않고도 바로 반응시킬 수 있습니다. 또한 TEV 반응물 자체를 Ni Spin Column에 그대로 로딩 해도 무방합니다. 단백질 구조와 정제 흐름 요약 pMAL-c6T 벡터를 사용하는 경우 단백질 구성은 다음과 같습니다: HHHHHH-MBP-linker (TEV protease 절단 서열 포함)-표적 단백질 이 구조의 단백질을 amylose resin에서 정제한 후 , His-tag를 갖는 TEV protease로 처리 하면 다음과 같은 결과물이 생성됩니다: HHHHHH-MBP-linker (절단 서열 포함) 표적 단백질 이 혼합물을 Ni Spin Column에 로딩하면: His-tag를 가진 TEV protease와 MBP-linker 는 Ni resin에 결합 표적 단백질은 flow-through로 분리 ...

Can Vaccinia Capping Enzyme Work on Short RNAs? Experimental Insights and Cleanup Strategies for 15nt–27nt Transcripts Experimental Insights and Cleanup Strategies for 15nt–27nt Transcripts When working with synthetic or transcribed RNAs, one common question arises: can very short RNA molecules—like 15 to 27 nucleotides—be efficiently capped using Vaccinia Capping Enzyme ? Although the literature and most protocols mention a minimum of 25nt, recent data and internal experiments suggest there’s more flexibility than previously thought. What’s the Minimum Length Required for Vaccinia Capping Enzyme? Traditionally, it's accepted that at least 25nt of RNA is needed for the enzyme to bind effectively and catalyze the addition of a 5′ cap ( Luo & Shuman, 1993 ). NEB’s internal tests, however, showed successful capping of 27mer RNAs , and even some 23nt RNAs in published studies reacted successfully. Th...

A Comprehensive Guide to Protein Quantitation Methods for Biopharmaceuticals A Comprehensive Guide to Protein Quantitation Methods In biopharmaceutical development, accurate protein quantitation is not just a routine step—it’s a fundamental requirement for assessing potency, purity, and stability. However, the choice of assay method can dramatically affect the reliability of your results, especially when dealing with chemically modified proteins such as PEGylated or glycosylated forms. Why Protein Quantitation Accuracy Matters According to ICH guidelines, protein quantitation directly influences structural characterization (like CD spectra), bioactivity measurement, and release criteria. Misjudged concentrations may lead to misinterpretation of assays or complete batch failures. Comparison of Six Major Protein Quantitation Assays Assay Principle ...

Hello everyone! Today, we’re going to learn how to introduce your desired mutations into plasmids easily using the Q5 Site-Directed Mutagenesis (SDM) Kit. SDM is an essential technique for studying gene function or protein structure–function relationships, and the Q5 Kit lets you perform what can be a complex process in a very streamlined way. Substitutions, Deletions, and Insertions — All Possible Key Features: Supports Substitutions , Deletions , and Insertions Optimized for small plasmids , ensuring high efficiency KLD reaction (Kinase, Ligase, DpnI) for one-tube circularization The Q5 SDM Kit is used to insert , swap , or remove specific nucleotide sequences in a plasmid. It leverages the KLD reaction to achieve both convenient circularization and removal of the original template in a single tube. SDM Primer Design Strategies 1. Overlapping Primer Design Principle: Primers extend around the plasmid and introduce a nick that promotes spontaneous circularizatio...