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As example, a local BlastN using the conserved domain V of RmInt1 (GAGCGGTGTGAATCGAGAGGTTTACGCACCGTTC) on these 6–8 kb can identify the potential domain V of the RNA structure. ucalgary. html) can be used to delimit the borders of the putative group II intron [9, 10]. In a canonical group II intron a “GUGYG” and “AY” as boundaries of 5′ and 3′ end, respectively, can be confirmed using this tool. q = mfold;) for the detection of potential group II intron RNA structures can be performed in order to identify the remaining RNA domains (I, II, III, and VI).

The OCT4 binding sites are mapped to hg18. First convert the coordinates to hg19 using LiftOver. chain. Extract repeat files. Format repeat data into a BED file. See http://bedtools. html for more information on this format. (a) One way to format is using an awk command. This example assumes this is the full RepeatMasker file from UCSC. bed. 1 requires that a minimum of 10 % of each repeat overlaps a binding site. (d) -u writes each repeat instance once if any overlaps are found. Use the BEDTools program shuffleBed to randomly permute the OCT4 binding regions in the genome for use as a control (see Note 1).

Sometimes it is useful to investigate if this group II intron is interrupting a particular gene. A BlastX using the corresponding DNA fragment lacking the proposed group II intron sequence can reveal a new copy of the interrupted ORF but in this case without the group II intron. This fact is also indicative that our sequence contains a full-length mobile group II intron. 2 Group II Intron Splicing The excision of group II introns can be detected by primer extension [11], and more accurately by RT-qPCR [12].

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