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Thus, calcium transients were defined as fluorescence signals (F/F0) that exceed 3.5 standard deviations above baseline, which were confirmed by frame-by-frame analysis of the time-lapse images.
The presence of a normal set of cells in the right cluster was confirmed by immunodetection (frame 9, right cluster).
This was confirmed by immuno-detection (frame 9) showing that the left cluster was formed by four outer cells: two Su(H) positive socket cells (yellow cells) and two shaft cells (green sibling cells), showing that the organ originated from two pIIa cells.
One of these alleles, dRecQ414, was further confirmed by DNA sequencing (Figure 1B) revealing that the start codon mutation and open reading frame shift are as designed.
Correct orientation and reading frame were confirmed by sequencing.
The DNA sequence in frame was confirmed by sequencing after transformation into DH5α competent cells (Invitrogen).
The start and the stop codons of each open reading frame were confirmed by comparison with homologous proteins.
The translational start and the stop of each open reading frame were confirmed by comparison with homologous proteins.
Sequences were assembled by multiple fragments from RACE and full length open reading frame was confirmed by PCR amplification and sequencing.
This open reading frame was confirmed by the genomic sequences of putative orthologues of this gene from P. glauca and P. abies.
The CD44st gene sequence and open reading frame were confirmed by restriction enzyme analysis and nucleotide sequencing, and then inserted into the eukaryotic expression vector pcDNA3.1.
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