Site Directed Mutagenesis (SDM) Rat

Plasmid isolation is an important technique in molecular biology or any kind of genetic editing. It involves amplifying plasmids overnight by transforming them into competent bacterial cells. The desired colonies of these bacteria can then be grown in shaker cultures, at appropriate shaking speed, oxygen availability and temperature. These liquid cultures can then be ultracentrifuged to pellet the bacteria, which are then used for plasmid isolation. The bacteria are first resuspended in a buffer, then lysed, neutralized, purified in a column, eluted, precipitated with ethanol and then resuspended. During plasmid isolation, it is important to lyse cells quickly because lysing bacteria for too long may lead to irreversible denaturing of the plasmid. Usually, alkaline lysis is used for isolation because it is a mild treatment. It isolates plasmid DNA and other cell components such as proteins by breaking cells apart with an alkaline solution. Precipitation removes the proteins, and the plasmid DNA recovers with alcohol precipitation. Resuspension and lysis buffers should be mixed thoroughly in order to prevent the DNA from breaking into smaller fragments. This is because broken gDNA can reanneal and remain in the solution, without binding to the column.

Get tips on using BH50bp ladder :: GD 50bp DNA Ladder RTU Ladder to perform DNA Ladder 50 bp

Get tips on using QuantiNova SYBR Green RT-PCR Kit (2500) to perform PCR Quantitative real-time PCR - Viral

Get tips on using QuantiFast Pathogen RT-PCR +IC Kit (400) to perform PCR Quantitative real-time PCR - Viral

Get tips on using QuantiTect SYBR Green RT-PCR Kit (1000) to perform PCR Conventional / Qualitative PCR - mammalian DNA

Get tips on using iScript™ Reverse Transcription Supermix for RT-qPCR to perform cDNA synthesis Cell lines

Get tips on using Superscript reverse tran-scriptase II (SS RT II) system to perform cDNA synthesis Yeast

Get tips on using BH100bp ladder :: GD 100bp DNA Ladder H3 RTU Ladder to perform DNA Ladder 100 bp

Get tips on using QIAGEN OneStep Ahead RT-PCR Kit (200) to perform PCR Quantitative real-time PCR - Fish species DNA

RNA-Seq is a method to sequence RNA by applying Next Generation Sequencing (NGS). The quality of RNA is critical for the success of RNA-Seq. The integrity of RNA is measured by the RNA integrity number (RIN). RIN is computed from RNA electrophoresis and electropherogram profiles (the peak area of the 28S rRNA should be approximately twice the peak area of the 18S rRNA). If you get the RIN value lower than 7, the possibility of getting the low quality of RNA-seq data is high. To get a high quality RNA, it is better to work with fresh samples or snap-freeze the tissues in liquid nitrogen as quickly as possible and store them at -80°C until further use. Make sure designated areas and all your filter tips, microfuge tubes, plastic, and glassware are RNase-free.