The folder "NN_frequency" contains the following files: 1. NN_frequency: a tab delimited .txt file generated by Perl software "thermo_NN'. The file contains two columns: - column 1: position of the second nucleotide in the nearest-neighbor(NN) couple - column 2: digits from 1 to 16 which represent a definite nearest-neighbor interaction in RNA/DNA duplex (see table 1 below ), where RNA is identical to Watson coding strand: Table 1 rAA/dTT=1 rUU/dAA=2 rAU/dTA=3 rUA/dAT=4 rCA/dGT=5 rUG/dAC=6 rGU/dCA=7 rAC/dTG=8 rCU/dGA=9 rAG/dTC=10 rGA/dCT=11 rUC/dAG=12 rCG/dGC=13 rGC/dCG=14 rGG/dCC=15 rCC/dGG=16 2.Access databases that contain "NN" in their names: these databases are designed to calculate the number of NN interactions in a given feature (genes,IGRs, introns). The Access databases: contain: 1.Tables: 1.1. Delta: an empty table in which the .txt file (see above) should be imported.This table is used by the module "Transform" to generate table "DeltaG" The table contains two columns, named: -position_bp: position of the second nucleotide in the nearest-neighbor(NN) couple -DeltaG: digits from 1 to 16 which represent a definite nearest-neighbor interaction in RNA/DNA duplex, where RNA is identical to Watson coding strand 1.2.Genes:in spite of its name this table is filled with information about a given feature (genes, IGRs, introns). This table is used by the module "Transform" to generate table "DeltaG" The table contains two columns, named: -ORF: this column contains the name or SGD number of a particular feature -position_start:coordinates of features' starts -position_end:coordinates of features' ends -strand: this column indicates the strand to which the definite feature belongs (Watson or Crick coding strand, indicated respectively by "W" and "C") Note: In NN_IGR there are three tables containing "Genes" in their name: "Genes" :an empty table , presented in the database in order to preserve the structures of the queries; "Genes_W": contains iformation about the coordinates of the IGRs and UTRs 3' to Watson's ORFs; "Genes_C": contains iformation about the coordinates of the IGRs and UTRs 3' to Crick's ORFs; To calculate NN frequence of one of these features (3' to Watson's or 3'to Crick's ORFs) you should delete "Genes" and rename "Genes_w" or "Genes_C" to "Genes" 1.3. DeltaG: an empty accessory table that should be filled by running the Module "Transform". Then the table would be filled with the following data: the strand to which the definite feature belongs in column "ORF" and the position of each nucleotide within the borders of the feature in column "position_bp". Note: All the tables should be filled with data before running the Queries 2.Queries: 2.1. Nucleotide: an accessory query, necessary for runnin query "Measurement". To measure the frequency of a particular NN interaction, open query "Nucleotide" in design view and in column "DeltaG" , row "Criteria" type the digit corresponding to particular NN interaction (for example: type 1 when measuring rAA/dTT frequency). 2.2. Measurement: generates a table containing the columns: - strand: "W" and "C" strand - CountOFDeltaG: the number of particular NN interaction in the investigated feature that belongs to either W or C strand. Note: A.For the Features belonging to "W" strand the number of digits obtained by query "Measurement" will correspond to the number of the particular NN interaction in the sense mRNA/DNA duplex if the NN interactions are indicated by the following digits: rAA/dTT=1 rUU/dAA=2 rAU/dTA=3 rUA/dAT=4 rCA/dGT=5 rUG/dAC=6 rGU/dCA=7 rAC/dTG=8 rCU/dGA=9 rAG/dTC=10 rGA/dCT=11 rUC/dAG=12 rCG/dGC=13 rGC/dCG=14 rGG/dCC=15 rCC/dGG=16 B.For the Features belonging to "W" strand the number of digits obtained by query "Measurement" will correspond to the number of the particular NN interaction in the antisense mRNA/DNA duplex if the NN interactions are indicated by the following digits: rUU/dAA=1 rAA/dTT=2 rUA/dAT=3 rAU/dTA=4 rUG/dAC=5 rCA/dGT=6 rAC/dTG=7 rGU/dCA=8 rAG/dTC=9 rCU/dGA=10 rUC/dAG=11 rGA/dCT=12 rGC/dCG=13 rCG/dGC=14 rCC/dGG=15 rGG/dCC=16 C.For the Features belonging to "C" strand the number of digits obtained by query "Measurement" will correspond to the number of the particular NN interaction in the sense mRNA/DNA duplex if the NN interactions are indicated by the following digits: rUU/dAA=1 rAA/dTT=2 rUA/dAT=3 rAU/dTA=4 rUG/dAC=5 rCA/dGT=6 rAC/dTG=7 rGU/dCA=8 rAG/dTC=9 rCU/dGA=10 rUC/dAG=11 rGA/dCT=12 rGC/dCG=13 rCG/dGC=14 rCC/dGG=15 rGG/dCC=16 D.For the Features belonging to "C" strand the number of digits obtained by query "Measurement" will correspond to the number of the particular NN interaction in the antisense mRNA/DNA duplex if the NN interactions are indicated by the following digits: rAA/dTT=1 rUU/dAA=2 rAU/dTA=3 rUA/dAT=4 rCA/dGT=5 rUG/dAC=6 rGU/dCA=7 rAC/dTG=8 rCU/dGA=9 rAG/dTC=10 rGA/dCT=11 rUC/dAG=12 rCG/dGC=13 rGC/dCG=14 rGG/dCC=15 rCC/dGG=16 Example: If you investigate the number of rAA/dTT in the genes you have to: 1. open database "NN_genes.mdb" 2. import "NN_frequency.txt" in table "Delta" 3. run the module "Transform" that will fill with data table "DeltaG" 4. open query "Nucleotide" in design view and in column "DeltaG" , row "Criteria" type the digit "1". (Table 1) 5. Run query "Measurement" You will obtain a table containing two values 1000000 for "W" and 700000 for "C" strand. This means that the genes belonging to "W" strand contain 1000000 rAA/dTT interactions in the sense mRNA/DNA duplex (Note A) and respectively 1000000 rUU/dAA interactions in the antisense RNA/DNA duplex (Note B).This is because these two RNA/DNA dinucleotide duplexes contain complementary DNA strands. Furthermore,the genes belonging to "C" strand contain 700000 rUU/dAA interactions in the sense mRNA/DNA duplex (Note C) and respectively 700000 rAA/dTT interactions in the antisense RNA/DNA duplex (Note D). To calculate the number of rAA/dTT in the genes belonging to "C" strand in sense mRNA/DNA duplexes, go back to procedure "4" and type digit "2" (Table 1). In that case you will obtain a table containing again two values 900000 for "W" and 800000 for "C" strand. This means that the genes belonging to "W" strand contain 900000 rUU/dAA interactions in the sense mRNA/DNA duplex (Note A) and respectively 900000 rAA/dTT interactions in the antisense RNA/DNA duplex (Note B). Furthermore,the genes belonging to "C" strand contain 800000 rAA/dTT interactions in the sense mRNA/DNA duplex (Note C) and respectively 800000 rUU/dAA interactions in the antisense RNA/DNA duplex (Note D).