Vectors for cloning

There exist a variety of vectors for cloning into eukaryotic systems, ranging from yeast (Saccharomyces as well as Pichia) through insect cells (Baculovims) and plants (Ti plasmid from Agrobacterium tumefaciens) to mammalian cells (transfected by viral or mammalian vectors). As expression in eukaryotic hosts is less efficient than bacterial expression in terms of yield and time and more complicated in terms of vector structure and culture conditions, such eukaryotic expression systems are only used for genes whose proteins require posttranslational modification which is not possible in bacteria. Yeast is the preferred option as a relatively easily culturable single-cell system but posttranslational modification capabilities is limited. The additional complexity can be circumvented in part by exploiting the ability of eukaryotic vectors to act as shuttle vectors, which can be shuttled between two evolutionarily different hosts. Thus, eukaryotic vectors can be replicated and analyzed in bacteria and transfected into eukaryotic cells for expression of the recombinant product. 

Phage Display Vectors for Cloning of Antibody Genes in Alphabetical Order co... 

Figure 9.8. Construction of cloning vector with BioEdit. The plasmid pJRD158 is retrieved from Entrez and used to construct vector for cloning DNA encoding human somatostatin with BioEdit. The cloning vector with somatostatin gene (arrow) is displayed.

Stoker, N. G., Fairweather, N. F., and Spratt, B. G. (1982). Versatile Low-Copy-Number Plasmid Vectors for Cloning in Escherichia coli. Gene 18 335. 

Development of safe and effective vectors for cloning the gene and inserting it into palients cells,... 

W, and Sahm, H. (1991) A family of Corynehacterium glutam-icum/Escherichia coli shuttle vectors for cloning, controlled gene expression, and promoter probing. Gene, 102 (1), 93-98. 

The product of the amplification may contain several bands that migrate at the same position in the sequencing gel. The best way to assess the identity of the hands in the first place is to clone the PCR product into a suitable vector for cloning PCR products. Several clones should be sequenced to verify homogeneity of the initial product. Once a major clone has been identified. Northern hlot hybridization should be performed using the cloned major PCR product as a probe. We do not recommend either dot-blot or Southern blot hybridization of the PCR products with a hot cDNA probe to verify positive clones, since repetitive sequences and labeled ribosomal RNA can give rise to nonspecific positive signals. 

Larson JL. Hershberger CL. Shuttle vectors for cloning recombinant DNA irt Eschericfiia colt and Streptomyces grtseo/liscus C581. J Bacteriol 1984 157 314-317. 

Rao RN, Richardson MA, Kuhstoss 5- Cosmid shuttle vectors for cloning and analysis of Streptomyces DNA. Meth Enzymol 1987 153 166-198. 

Breeding of Vectors, (a) Plasmids. Plasmids continue to be one of the major classes of vector for cloning and amplification of DNA molecules. The most commonly used plasmids of the mid-to-late 1970 s, pSClOl and ColEl, have been modified to make them more effective as cloning vehicles. 

Figure 2 A scheme for the terminal addition of restriction etiAo-nuclease EcoRl recognition sites to large fragments q/ DNA. The products can be inserted into the EcoRl site of a vector for cloning...

Godiska R, Mead D, Dhodda V, Wu C, Hochstein R, Karsi A, Usdin K, Entezam A, Ravin N. (2010). Linear plasmid vector for cloning of repetitive or unstable sequences in Escherichia coli. Nucleic Acids Research.YoL 38(6) e88. 

Pierce J.C., Sauer B., and Sternberg N. 1992. A positive selection vector for cloning high molecular weight DNA by the bacteriophage PI system Improved cloning efficacy. Proc. Natl. Acad. Sci. 89 2056-2060. 

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