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construction and characterization of a cdna library from wheat infected with fusarium graminearum fg 2建设和表征的cdna文库花药对赤霉病的小麦感染fg 2.pdf

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construction and characterization of a cdna library from wheat infected with fusarium graminearum fg 2建设和表征的cdna文库花药对赤霉病的小麦感染fg 2

Int. J. Mol. Sci. 2011, 12, 613-626; doi:10.3390/ijm OPEN ACCESS International Journal of Molecular Sciences ISSN 1422-0067 /journal/ijms Article Construction and Characterization of a cDNA Library from Wheat Infected with Fusarium graminearum Fg 2 Khaled Al-Taweel 1,2, W. G. Dilantha Fernando 1,* and Anita L. Brûlé-Babel 1 1 Department of Plant Science, University of Manitoba, Winnipeg, Manitoba R3T 2N2, Canada; E-Mails: khaledta72@ (K.A.-T.); anita_brulebabel@umanitoba.ca (A.B.-B.) 2 General Commission of Agricultural Scientific Research (GCASR), Department of Biotechnology, P.O. Box 113 Douma-Damascus, Syria * Author to whom correspondence should be addressed; E-Mail: D_Fernando@umanitoba.ca; Tel.: 204-474-6072; Fax: 204-474-7528. Received: 26 November 2010; in revised form: 14 January 2011 / Accepted: 17 January 2011 / Published: 18 January 2011 Abstract: Total RNA from wheat spikes infected with F. graminearum Fg2 was extracted and the mRNA was purified. Switching Mechanism at 5 end of the RNA Transcript (SMART) technique and CDS Ill/3 primer were used for first-strand cDNA synthesis using reverse transcriptase by RT-PCR. Primer extension polymerase chain reaction was used to construct the double-strand cDNA that was digested by proteinase K, then by Sfi I and fractionated. cDNAs longer than 0.5 kb were collected and ligated to λTriplEx2 vector followe