인천대학교 나노바이오실험(1) A 자료) 4. Plasmid perp & Gel Electrophoresis

문서 내 토픽

1. Plasmid DNA

플라스미드 DNA는 세균의 세포 내에 염색체와는 별개로 존재하면서 독자적으로 증식할 수 있는 DNA이다. 유전공학에서는 세균 내 플라스미드를 세포 밖으로 빼내고 제한효소로 끊은 뒤, 필요로 하는 유전자를 삽입하여 이를 다시 세균에 넣어 배양하는 유전자재조합 기술을 사용한다.
2. Plasmid DNA 추출 방법

플라스미드 DNA를 추출하는 방법에는 크기 차이를 이용한 방법과 구조 차이를 이용한 알칼리 용해 방법(Alkaline Lysis method)이 있다. 알칼리 용해 방법은 높은 pH의 알칼리성 용액을 이용하여 대장균 세포를 용해시키고 플라스미드 DNA를 염색체 DNA와 구분하여 분리하는 실험방법이다.
3. Gel Electrophoresis

전기영동은 핵산을 식별, 정량화 및 정제하는 데 사용되는 일반적인 실험기술이다. DNA, RNA, 단백질과 같이 전하를 띤 분자를 분리하기 위해 흔히 사용된다. 모든 DNA 분자는 질량 당 동일한 양의 전하를 갖기 때문에, 크기에 따라서 분리된다. 아가로스 겔을 이용한 수평 전기영동 방식이 DNA 분리에 주로 사용된다.
4. Nanodrop

Nanodrop은 DNA, RNA, 단백질의 아주 적은 소량 정량 측정에 적합하도록 고안된 장비로, 전 파장의 흡광도를 동시에 짧은 시간만을 소모 측정하여 Real Scanning으로 Sample의 상태를 보다 쉽고 간편하게 확인할 수 있다.

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1. Plasmid DNA

Plasmid DNA is a circular, double-stranded DNA molecule that is found in bacteria and some eukaryotic cells. It is distinct from the linear chromosomal DNA and is capable of autonomous replication within the host cell. Plasmids often carry genes that provide the host cell with beneficial traits, such as antibiotic resistance or the ability to metabolize certain compounds. They are widely used in molecular biology and biotechnology as vectors for gene cloning, expression, and manipulation. Plasmid DNA is a crucial tool in genetic engineering, allowing researchers to introduce foreign genes into host cells and study their function or produce valuable proteins. Understanding the structure, properties, and applications of plasmid DNA is essential for advancements in fields like medicine, agriculture, and environmental science.
2. Plasmid DNA 추출 방법

Plasmid DNA extraction is a fundamental technique in molecular biology and biotechnology. There are several methods available, each with its own advantages and disadvantages. The most common methods include alkaline lysis, boiling, and commercial kits. Alkaline lysis involves breaking down the bacterial cell wall and membrane, followed by neutralization and precipitation of the plasmid DNA. Boiling is a simpler method that relies on the heat-induced denaturation of cellular components, allowing the plasmid DNA to be separated. Commercial kits, on the other hand, utilize specialized resins or columns to selectively bind and purify the plasmid DNA, often providing higher purity and yield. The choice of extraction method depends on factors such as the size and copy number of the plasmid, the desired purity, and the downstream applications. Efficient and reliable plasmid DNA extraction is crucial for various applications, including gene cloning, protein expression, and genetic engineering.
3. Gel Electrophoresis

Gel electrophoresis is a powerful analytical technique used in molecular biology and biochemistry to separate and analyze DNA, RNA, or proteins based on their size and charge. It involves the migration of charged molecules through a gel matrix under the influence of an electric field. This method allows for the separation and visualization of different DNA fragments, plasmids, or protein samples, enabling researchers to study their size, purity, and relative abundance. Gel electrophoresis is widely used in various applications, such as DNA sequencing, gene expression analysis, protein profiling, and forensic DNA fingerprinting. The technique provides valuable information about the molecular composition of samples and is an essential tool in fields like genetics, molecular biology, and biotechnology. Continuous advancements in gel electrophoresis, including the development of automated systems and specialized gels, have further expanded its capabilities and applications in scientific research and diagnostic procedures.
4. Nanodrop

The Nanodrop is a spectrophotometric instrument used for the quantification and purity assessment of nucleic acids (DNA, RNA) and proteins. It is a valuable tool in molecular biology and biochemistry laboratories, as it allows for the rapid and accurate measurement of sample concentrations and quality. The Nanodrop utilizes a small sample volume (typically 1-2 μL) to determine the absorbance of the sample at specific wavelengths, which can then be used to calculate the concentration of the target molecule. This is particularly useful for analyzing precious or limited sample sizes, as it minimizes the amount of material required for analysis. Additionally, the Nanodrop provides information on the purity of the sample by calculating the A260/A280 and A260/A230 ratios, which indicate the presence of contaminants such as proteins, phenols, or salts. The Nanodrop's ease of use, speed, and versatility make it an indispensable tool in various fields, including molecular biology, biochemistry, and biotechnology, where accurate and efficient quantification and quality assessment of nucleic acids and proteins are crucial for experimental success and data reliability.