Representative cDNA libraries from few plant cells.

T Dresselhaus; H Lörz; E Kranz

doi:10.1046/j.1365-313x.1994.5040605.x

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Representative cDNA libraries from few plant cells.

T Dresselhaus·H Lörz·E Kranz

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Abstract

A reverse transcriptase/polymerase chain reaction (RT/PCR) method for the construction of representative cDNA libraries originating from few isolated cells is described. Poly(A)+ RNA was extracted from an average of 100 maize cells and reverse transcribed into sscDNA. The sscDNA was dG-tailed at its 3' end and amplified during a two-step PCR reaction. The generated PCR products were analysed and the majority < or = 2 kbp were full-size cDNAs. A fraction of the amplified cDNA from 128 isolated maize egg cells was cloned into the lambda Uni-ZAP XR vector and a primary library of 6.8 x 10(6) p.f.u. was obtained. The average insert size is 860 bp. It was further determined, that 0.31% of the clones hybridized to a cytosolic GAPDH probe. It is thought that, with this method, the first cDNA library of egg cells in higher plants was generated.

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Base Sequence DNA Primers DNA, Complementary Gene Library Genetic Techniques Molecular Sequence Data Oligodeoxyribonucleotides Polymerase Chain Reaction Protoplasts Zea mays Journal Article Research Support, Non-U.S. Gov't

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T Dresselhaus, H Lörz, E Kranz. 1994. Representative cDNA libraries from few plant cells.. https://doi.org/10.1046/j.1365-313x.1994.5040605.x

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