Experimental Analysis of XPCR-based protocols

This paper reports some experimental results validating in a broader context a variant of PCR, called XPCR, previously introduced and tested on relatively short synthetic DNA sequences. Basic XPCR technique confirmed to work as expected, to concatenate two genes of different lengths, while a library of all permutations of three different genes (extracted from the bacterial strain Bulkolderia fungorum DBT1) has been realized in one step by multiple XPCR. Limits and potentialities of the protocols have been discussed, and tested in several experimental conditions, by aside showing that overlap concatenation of multiple copies of one only gene is not realizable by these procedures, due to strand displacement phenomena. In this case, in fact, one copy of the gene is obtained as a unique amplification product.

[1]  Takahiro Kanagawa,et al.  Bias and artifacts in multitemplate polymerase chain reactions (PCR). , 2003, Journal of bioscience and bioengineering.

[2]  Karl-Heinz Zimmermann,et al.  DNA Computing Models , 2008 .

[3]  T. Ceska,et al.  Structure-specific DNA cleavage by 5' nucleases. , 1998, Trends in biochemical sciences.

[4]  Daniel J. Park.  PCR Protocols , 2011, Methods in Molecular Biology.

[5]  Ehud Shapiro,et al.  Processing DNA molecules as text , 2010, Systems and Synthetic Biology.

[6]  Dario Neri,et al.  DNA-encoded chemical libraries: foundations and applications in lead discovery. , 2016, Drug discovery today.

[7]  Mohan Krishnamoorthy,et al.  A multiple-alignment based primer design algorithm for genetically highly variable DNA targets , 2013, BMC Bioinformatics.

[8]  T. Head.  Formal language theory and DNA: an analysis of the generative capacity of specific recombinant behaviors. , 1987, Bulletin of mathematical biology.

[9]  Vincenzo Manca,et al.  Algorithmic applications of XPCR , 2011, Natural Computing.

[10]  Giuditta Franco,et al.  A Polymerase Based Algorithm for SAT , 2005, ICTCS.

[11]  J. Dahlberg,et al.  Structure-specific endonucleolytic cleavage of nucleic acids by eubacterial DNA polymerases. , 1993, Science.

[12]  Vincenzo Manca,et al.  Computing by polymerase chain reaction. , 2008, Mathematical biosciences.

[13]  N. V. Pavlova,et al.  Recent developments in the optimization of thermostable DNA polymerases for efficient applications. , 2004, Trends in biotechnology.

[14]  R. Abramson,et al.  Detection of specific polymerase chain reaction product by utilizing the 5'----3' exonuclease activity of Thermus aquaticus DNA polymerase. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[15]  Carlo Laudanna,et al.  DNA Recombination by XPCR , 2005, DNA.