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. Author manuscript; available in PMC: 2010 Jan 1.
Published in final edited form as: Mol Microbiol. 2008 Nov 4;71(2):273–290. doi: 10.1111/j.1365-2958.2008.06530.x

Fig. 4. RLM-RACE analysis of esp transcripts.

Fig. 4

A. Agarose gel electrophoresis of the products obtained after RLM-RACE. esp transcripts were analyzed by 5’-RLM-RACE employing as a substrate a RNA sample extracted from a DMEM-grown culture of Sakai. The RNA sample was treated with calf intestine phosphatase (CIP) and tobacco acid pyrophosphatase (TAP) in different combinations to detect primary (Pri) and processed (Pro) transcripts (see Experimental Procedures). After the indicated treatments and reverse transcription, nested PCR was performed with 5’-RACE and espA specific primers (Table I). Lanes 1, 3, 5 and 7 correspond to the first PCR and lanes 2, 4, 6, and 8 correspond to the second nested PCR. (M) 1 kb Plus DNA Ladder (Invitrogen)

B. 5’ end locations of ‘esp’ transcripts. The products obtained after 5’-RLM-RACE were cloned and sequenced (reactions shown in lanes 2 and 8 in (A)). The 5’ ends are shown in capital letters and their distribution in 47 clones is indicated by a large arrow (18 clones), small arrow (11 clones), underlined nucleotides (5 clones each nucleotide), and double underlined nucleotides (4 clones each nucleotide). The horizontal arrows indicate the region analyzed by RPA.