Construction of cDNA Libraries
To obtain a cDNA clone of an mRNA, the mRNA must be copied faithfully into DNA, and the cDNA library must be large enough to represent the abundance class that contains the mRNA of interest. For example, in tobacco, Goldberg (1 ) has shown that it is possible to divide the mRNA population into three classes with most of the mRNAs (11,300) being in the lowest abundance class and making up 39% of the polysomal rnRlVA. To obtain a cDNA library that contains at least one clone for each mRNA of this class will require about 2 � 105 clones (2 ). This can be achieved with a few micrograms of mRNA by using the efficient RNase H method of making double-stranded cDNA (3 ; see Chapter 24 ) and a bacteriophage λ vector that exploits the high efficiency with which in vitro packaged phage can be introduced into Escherichia coli . The choice of λ vector is important because if the DNA to be inserted makes the λ genome > 105% of the wild-type length, the packaged phage will have a low viability.
- 生物发光和化学发光(BL/CL)用于基因表达
- Detection of Copy Number Changes at Multiple Loci in DNA Prepared from Formalin-Fixed, Paraffin-Embedded Tissue by Multiplex Lig
- HAT: A Novel Statistical Approach to Discover Functional Regions in the Genome
- Construction of Oligonucleotide Microarrays (Biochip) Using Heterobifunctional Reagents
- Identification of Messenger RNAs and MicroRNAs Associated With Fragile X Mental Retardation Protein
- Ultrasensitive Detection of DNA Damage by the Combination of the Comet and TUNEL Assays
- QTL Analysis in Plants
- The Detection of Latent Virus Infection by Polymerase Chain Reaction
- Electron Microscopy of Polytene Chromosomes
- Cycle Sequencing of PCR Products