Cloning and sequencing HAR1 and NTS1
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چکیده
A 72 kb region of BAC259.12D that encompasses the HAR1 locus was sequenced in its entirety using a DNA Sequencing Kit (PE Applied Biosystems) with an automated DNA sequencer (ABI PRISM 3100; PE Applied Biosystems). HAR1 complementary DNA was cloned from a cDNA library of L. japonicus shoots using DNA fragments of the first exon of the HAR1 gene obtained by polymerase chain reaction (PCR) from BAC259-12D. After sequencing the cDNA clone, the full-length cDNA sequence was determined by 5 0 rapid amplification of cloned ends (RACE) on L. japonicus shoot messenger RNA. The soybean gene showing the highest similarity with HAR1, G. max CLV1B, was amplified from the genomic DNA of a soybean cultivar Enrei and of a hypernodulating mutant, En6500, by PCR with a specific primer set designed from the DNA Data Bank of Japan (DDBJ) database. They were sequenced using a DNA Sequencing Kit (PE Applied Biosystems) with an automated DNA sequencer.
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تاریخ انتشار 2002