RNA binding strengths of centromeres and the flanking sequences on 19 and X chromosomes

A total of 1,000 genes highly expressed in human tonsil germinal center B cells were selected for analysis based on the results of the Digital Differential Display (NCBI UniGene Lib.289 -NCI_CGAP_GCB1). Gene-Analyser 2.0 software (see Gene-Analyser 2.0) was used to analyze the number of 7-nucleotide (7-nt) strings (47=16,384), which was written by our team. The 50-kb DNA sequence was represented by a long column of numbers whose sum was 49,994.

Microsoft Excel software 2003 was used for statistical analysis.The algorithm for the RNA binding strengthThe RNA binding strength algorithm is based on the principle that more complementarity between RNA and DNA results in more binding between RNAs and DNAs. For example, when there is one 5'-TTTTTTT DNA molecule and ten 5'-AAAAAAA RNA molecules in a certain volume solution, the likelihood of DNA binding with RNAs is 10 (10×1=10).

If there are ten 5'-TTTTTTT DNA molecules, the likelihood of DNA binding with RNAs is 100 (10×10=100). The binding of single-strand RNA and double-strand DNA accounts for competition between RNA and DNA for binding.The centromere of chromosome 19 (chr19) is located at 24.50Mb-27.19Mb. DNA sequence from 19Mb to 32Mb of chromosome 19 was divided into different 50kb fragments.

Gene-Analyser 2.0 software was used to analyze the 7nt strings contained in each 50kb fragment. The results of analysis was shown in folder of Chr19 sequence and 7 nt strings. Folder, named “19”, contains the DNA base sequences and their 7nt strings from 19000001 bp to 20000000 bp of chr19 and contains 20 text files and 20 Excel files.

Folders 20-32 represent the same meaning as folder “19”.The centromere of chromosome X (chrX) is located at 58.61Mb-62.41Mb. DNA sequence from 52Mb to 67Mb of chrX was divided into different 50kb fragments. Gene-Analyzer 2.0 software was used to analyze the 7nt strings contained in each 50kb fragment.The results of analysis was shown in folder of ChrX sequence and 7 nt strings.

Folder 52 contains the DNA base sequences and their 7nt strings from 52000001 bp to 53000000 bp of chrX and contains 20 text files and 20 Excel files. Folders 53-67 represent the same meaning as folder 52.The total RNA of 1,000 genes highly expressed in tonsil germinal center B cells is shown in Excel file, named “7nt strings of RNA expressed by 1000 genes”.7nt string data of each 50kb fragement is multiplied by the total RNA of same strings, and the sum of 16,384 products is the RNA binding strength of the 50 kb fragement.The 50 kb fragment size was chosen because a transcription unit contains 10-50kb sequences (based on DNase I digestion of the hemoglobin and ovoalbumin genes).

One-thousand genes highly expressed in human tonsil germinal center B cells were selected (as described above), and the 7-nt string numbers for these genes were calculated from the sense strand (including introns and exons). The 7-nt string numbers for each gene multiply by the expression frequency of the gene (Lib.5601; ncbi.nlm.nih.gov/UniGene/), which results in the calculated numbers of the 7-nt string for the gene (see excel file, named “ChrX52 RNA binding strength”, shows how to calculate the RNA binding strength value of one 50-kb DNA fragment (from 52000001 bp to 53000000 bp of chrX ).Excel file, named “Chr19”, shows the result of RNA binding strengths of centromeres and the flanking sequences of Chr19.

The Excel file, named “ChrX” , shows the value of RNA binding strengths of centromeres and the flanking sequences of chrX.