scholarly journals Production of Bovine Transgenic Conceptus; Possible Selection of Transgenic Embryos by Polymerase Chain Reaction.

1999 ◽  
Vol 45 (6) ◽  
pp. 387-395
Author(s):  
Masaki Hoshi ◽  
Kazuhiro Saeki ◽  
Yoshikazu Nagao ◽  
Shu Hashimoto ◽  
Noboru Seike ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Transgenic Embryos ◽  
Selection Of

Efficient selection of preimplantation transgenic embryos by an improved procedure using Dpn I - Bal 31 digestion and the polymerase chain reaction

1997 ◽  
Vol 9 (2) ◽  
pp. 263 ◽  
Author(s):  
B. B. Seo ◽  
C.H Kim ◽  
H. Tojo ◽  
S. Tanaka ◽  
K. Yamanouchi ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Dna Sequence ◽  
Mouse Embryos ◽  
Chain Reaction ◽  
Exogenous Dna ◽  
Pcr Products ◽  
Polymerase Chain ◽  
Transgenic Embryos ◽  
Efficient Selection ◽  
Selection Of

Efficient selection of preimplantation transgenic embryos by an improved method after pronuclear injection of exogenous DNA is described. The method is based on subjecting DNA extracted from the embryos to restriction enzymes as well as the polymerase chain reaction (PCR). The incorporated procedure included recovery of the digested DNA with glassmilk before PCR, which markedly enhanced the rate of accurate detection of transgenic embyos. When exogenous DNA sequences in the mouse embryos were not integrated into the genome they were digested with both Dpn I and Bal31, and subsequent PCR analysis generated DNA fragments of the injected DNA sequence in only 1 ·5% of cases examined. However, DNA extracted from mouse embryos containing the transgene sequences integrated into the genome evaded digestion by both enzymes and yielded transgene-specific PCR products in 68· 6% of the embryos tested. When bovine embryos were used, sequences of the endogenous haemoglobin gene used as a control genomic DNA sequence were protected from enzyme digestion (PCR products in 70· 5% of the embryos examined); by contrast, the non-integrated injected sequences were almost completly eliminated by the same treatment (PCR products in 1· 4% of the embryos examined). It is suggested that this method might be useful for the selection of transgenic embryos before embryo transfer, thereby reducing the number of recipient females required.

scholarly journals Easy and Efficient DNA Extraction from Woody Plants for the Detection of Phytoplasmas by Polymerase Chain Reaction

Plant Disease ◽  
1999 ◽  
Vol 83 (5) ◽  
pp. 482-485 ◽  
Author(s):  
Margaret J. Green ◽  
Dan A. Thompson ◽  
Donald J. MacKenzie
Keyword(s):  
Polymerase Chain Reaction ◽  
Plant Material ◽  
Woody Plant ◽  
Leaf Roll ◽  
Chain Reaction ◽  
Efficient Procedure ◽  
Identical Manner ◽  
Total Dna ◽  
Polymerase Chain ◽  
Selection Of

A simple and efficient procedure for the extraction of high-quality DNA from phytoplasma-infected woody and herbaceous plants for polymerase chain reaction (PCR) detection is described. This procedure does not require phenol, chloroform, or alcohol for the precipitation of nucleic acids. Herbaceous and woody plant material are extracted in an identical manner with no additional purification or enrichment steps required. The method utilizes commercially available microspin-column matrices, and the extraction of total DNA can be achieved in less than 1 h. The method has been used to successfully purify phytoplasma DNA from whole leaves, leaf petioles and midribs, roots, and dormant wood from a diverse selection of plant material. The phytoplasmas detected by PCR include pear decline, western X-disease, peach yellow leaf roll, peach rosette, apple proliferation, Australian grapevine yellows, and Vaccinium witches'-broom.

The use of deficiencies to determine essential gene content in the let-56–unc-22 region of Caenorhabditis elegans

Genome ◽  
1993 ◽  
Vol 36 (6) ◽  
pp. 1148-1156 ◽  
Author(s):  
Jacquie E. Schein ◽  
Marco A. Marra ◽  
Guy M. Benian ◽  
Chris Fields ◽  
David L. Baillie
Keyword(s):  
Polymerase Chain Reaction ◽  
Caenorhabditis Elegans ◽  
Genomic Sequence ◽  
Essential Gene ◽  
Chain Reaction ◽  
Radiation Induced ◽  
Practical Tool ◽  
Polymerase Chain ◽  
Selection Of

We have investigated the possibility of using the polymerase chain reaction to detect deletions of coding elements in the unc-22–let-56 interval on chromosome IV in the nematode Caenorhabditis elegans. Our analysis of approximately 13 kb of genomic sequence immediately to the left of the unc-22 gene resulted in the identification of four possible genes. Partial cDNAs have been identified for three of them. To determine whether any of these coding elements are essential for development, we required a method for the induction and selection of mutations in these elements. Our approach was to identify a set of formaldehyde and gamma radiation induced unc-22 mutations that mapped to the unc-22–let-56 region, and then employ polymerase chain reaction methodology to identify deficiencies that affected one or more of the four identified coding elements. Two small deficiencies were identified in this manner. Characterization of these deficiencies shows that there are no coding elements between unc-22 and let-56 (the nearest mutationally identified gene to the left of unc-22), which are required in development under laboratory conditions. We conclude that the polymerase chain reaction is a practical tool for the detection of deletions of coding elements identified in this region, and that characterization of such deficiencies provides a method for assessing whether or not these elements are required for development.Key words: Caenorhabditis elegans, deficiencies, coding elements, unc-22.

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