help_features

Input CRE

Input cis regulatory elements in "CRE features analysis module" of AtREA can be presently in consensus format only. In addition to consensus sequences based the four basic nucleotides i.e A,C,G and T.The feature analysis module also accepts the following nucleotide codes. For analysis of "variants" the input CRE can contain only A.C,G and T.

Code	Implication
M	A or C
R	A or G
W	A or T
S	C or G
Y	C or T
K	G or T
V	A or C or G
H	A or C or T
D	A or G or T
B	C or G or T
N	G or A or T or C

Example of input CRE

In addition to user defined CREs known CRE sequences from different CRE databases like PLACE and ATCISDB can be also analysed using AtREA. input field.

Analysis

Four different features of a CRE in a user selected class can be analysed by the "CRE feature analysis module".

Strand: The strand analysis option compares the occurrence of the input CRE in different strands(coding reverse and both) in the selected class to to their occurrences(in the corresponding strand) in the reference set.

Position: The position analysis compares the occurrence of the input CRE in five position windows (0-200, 200-400, 400-600, 600-800, 800-1000) in the selected class to their occurrences(in the corresponding position windows) in the reference set.

Frequency: In frequency analysis, all possible frequency states of a CRE (number of instances which a CRE can show in any of the 1Kb upstream sequence from the reference set) are identified. The CRE RCCGAC for example can occur 0,1,2,3,4 or 5 times in a 1Kb upstream sequence. The program the compares the occurrence of each frequency state of the input CRE in the selected class to its occurrence in reference set .

Variants: In this analysis the program generates all possible variants (differeing from the input CRE at the selected nucleotide position) and compare occurrence of each variant in the selected class to its occurrence in reference set. For CREs which contain six or less nucleotides the variants anlaysis for all the nucleotide positions can be performed in a single run by using the "all" option from the "Variant Position" menu. For longer CREs the user needs to select any nucleotide position of the CRE from the "Variant Position" menu.

*The reference set consists of 22229 1Kb upstream sequences(with respect ot TSS).
*The enrichment of a CRE in all these cases is estimated on the basis of cumulative hypergeometric probablity of observing greater or same number of CRE containing genes (in comparison to the class) in a random gene set of same size.

Class

Any class from any of the five ontology categories can be analysed for the enrichment of the user provided CRE. To begin with user needs to select a category from the list of class categories.For each of these categories, the class of interest can be choosen from the drop down menu of the corresponding ontology group.

Category	Ontology	Classes Incuded in AtREA
GOBP	Gene Ontology Biological Process	list of GOBP classes
GOMF	Gene Onltology Molecular Function	list of GOMF classes
GOCC	Gene Onlolotgy Cellular Component	list of GOCC classes
MIPS	MIPS FUNCAT classes	list of MIPS FUNCAT classes
ARACYC	ARACYC pathways	list of ARACYC classes

For example to analyse the features of a CRE is GOBP class GO:0000038 the user needs to select GOBP from the "category menu" and the class GO:0000038 from the "GOBP class" menu.

Select Class Category:

GOBP class

To study the features of a CRE in induced or repressed genes from any of the expression slides the corresponding option should be selected from the category menu.

Category	Description	Expression slides
induced	Genes which show induction in different microarray experiemnts	slides
repressed	Genes which show induction in different microarray experiemnts	slides

The slide of interest can then be selected from the "expression slide" menu.

Expression slide

As the characterization of genes as induced on repressed depends on a expression score cutoff the user can specify expression cutoff from the "Fold Cutoff" menu. The options include score cutoffs in log ₂ format and range from 1.2 to 4 (i.e. ~1.4 to 16 folds).

Fold Cutoff (only for expression classes) :

User can also analyse different features of a CRE in their own set of genes.For this the category "UserGeneset" should be selected as category and a list of AGI codes( like At1g75820) for genes shold be entered in the UserGeneSet text input box.

Select Class Category:

User Geneset
AT1G09830 AT1G10700 AT4G02260 AT4G11610 AT4G12440 AT4G13720 AT4G13890 AT4G16265 AT4G17360 AT4G22570 AT4G30530 AT4G30540 AT4G30550 AT4G34740 AT4G34890 AT4G38200 AT4G38880 AT5G03300 AT5G06850 AT5G09290 AT5G11160 AT5G19040 AT5G48060 AT5G67520 AT5G48060 AT5G67520

Output

Class feature (strand)	Number of CRE containing genes in class(Gc)	Number of genes in class(Tc)	Number of CRE containing genes in reference set(Gr)	Number of genes in reference set(Tr)	Percentage of CRE containing gene is class (Pc = (Gc/Tc) X 100 )	Percentage of CRE containing genes in reference set ( Pr=(Gr/Tr) X 100 )	Ratio (Pc/Pr)	Hypergeometic P-value	Genes which contain input CRE
Coding	7	16	1901	22229	43.750	8.552	5.12	0.0001892	At1g68020,At2g22190, At4g12430,At4g22590, At4g39770,At5g51460, At5g65140

Figure 1

The output of AtREA CRE features module (Figure 1) contains the following columns

Column 1: The name of the feature subclass. (For position analysis the subclasses correspond to different position windows.For strand analysis the feature subclasses are coding ,reverse and both ie. different strands.For variants each variant is treated as a feature subclass.For frequency analysis each frequency state is considered as a subclass.)
Column 2: Number of genes from class which show the particular CRE feature.
Column 3: Number of genes in the class.
Column 4: Number of genes from the reference set which show the particular CRE features.
Column 5: Number of genes in the reference set .
Column 6: Percentage of promoters from class which show the CRE feature.
Column 7: Percentage of promoters from reference set which show the CRE. feature
Column 8 (Hypergeometric P-vlaue) : the cumulative hypergeometric P-values of observing eqaul or greater number of CRE containing promoters in a set containing same number of genes by chance.
Column 9 : list of genes from class which contain the user defined CRE in their upstream sequence.

Example

The dehydration response element binding (DREB) motif is known to be involved in transcription regulation under draught stress. We have used different analysis option to study the distribution of DREB consensus (RCCGAC) sequence in genes which are induced by draught stress(slide:drought_6h_shoot(1007966668#1029, fold cutoff: 1.4).

Example of Strand analysis option

Table 1 shows the distribution of DREB consensus (RCCGAC) sequence (in different strands) among genes which are induced by drought stress(slide: drought_6h_shoot(1007966668#1029) , fold cutoff: 1.4).

CRE Strand	Number of CRE containing genes in class(Gc)	Number of genes in class(Tc)	Number of CRE containing genes in reference set(Gr)	Number of genes in reference set(Tr)	Percentage of CRE containing gene is class( (Gc/Tc) X 100 )	Percentage of CRE containing genes in reference set ( (Gr/Tr) X 100 )	Hypergeometic P-value	Genes which contain input CRE
coding	37	185	2895	22229	20.0000	13.0235	0.00488504236088069	At1g06460,At1g27630,At1g48330,At1g49720,At1g69830,At2g01520,At2g15880,At2g15970,At2g17840,At2g21660,At2g22450,At2g39030,At2g42530,At2g42540,At3g10410,At3g15630,At3g22740,At3g46640,At3g50970,At3g59350,At3g63160,At4g09020,At4g12280,At4g19120,At4g27440,At4g33700,At4g35770,At5g06690,At5g07010,At5g15960,At5g25210,At5g48250,At5g50450,At5g52310,At5g57630,At5g62360,At5g63810
reverse	42	185	2723	22229	22.7027	12.2498	4.96347784419354e-05	At1g22370,At1g26665,At1g29395,At1g51090,At1g75190,At1g76590,At2g02990,At2g16660,At2g19450,At2g25930,At2g28900,At2g29630,At2g34930,At2g38465,At2g39030,At2g42530,At2g42540,At2g47890,At3g05880,At3g18080,At3g22740,At3g26580,At3g28290,At3g46640,At3g47860,At3g48360,At3g50970,At3g53990,At3g62550,At4g01130,At4g02370,At4g09020,At4g11310,At4g12280,At4g19120,At4g23600,At4g30650,At5g03240,At5g06690,At5g26340,At5g47240,At5g57110
both	69	185	5221	22229	37.2973	23.4873	1.64100850699939e-05	At1g06460,At1g22370,At1g26665,At1g27630,At1g29395,At1g48330,At1g49720,At1g51090,At1g69830,At1g75190,At1g76590,At2g01520,At2g02990,At2g15880,At2g15970,At2g16660,At2g17840,At2g19450,At2g21660,At2g22450,At2g25930,At2g28900,At2g29630,At2g34930,At2g38465,At2g39030,At2g42530,At2g42540,At2g47890,At3g05880,At3g10410,At3g15630,At3g18080,At3g22740,At3g26580,At3g28290,At3g46640,At3g47860,At3g48360,At3g50970,At3g53990,At3g59350,At3g62550,At3g63160,At4g01130,At4g02370,At4g09020,At4g11310,At4g12280,At4g19120,At4g23600,At4g27440,At4g30650,At4g33700,At4g35770,At5g03240,At5g06690,At5g07010,At5g15960,At5g25210,At5g26340,At5g47240,At5g48250,At5g50450,At5g52310,At5g57110,At5g57630,At5g62360,At5g63810

Table 1

Example of Position analysis option:

Table 2 shows the distribution of DREB consensus (RCCGAC) sequence (in different position windows) in genes which are induced by draught stress(slide: drought_6h_shoot(1007966668#1029) , fold cutoff: 1.4). The results show that the frequency of the DREB consensus sequence increases as we move towards the TSS ,with maximum frequency in the 800-1000 window,in the selected gene set as well as the reference set.

CRE Position

Number of CRE containing genes in class(Gc)

Number of genes in class(Tc)

Number of CRE containing genes in reference set(Gr)

Number of genes in reference set(Tr)

Percentage of CRE containing gene is class( (Gc/Tc) X 100 )

Percentage of CRE containing genes in reference set ( (Gr/Tr) X 100 )

Hypergeometic P-value

Genes which contain input CRE

0 - 200

185

1088

22229

9.1892

4.8945

0.00953949701390473

At1g27630,At1g49720,At2g15880,At2g19450,At2g29630,At2g39030,At3g18080,At3g22740,At3g46640,At3g48360,At3g53990,At3g63160,At4g01130,At4g09020,At4g27440,At5g57110,At5g63810

200 - 400

185

1038

22229

6.4865

4.6696

0.157457096695083

At1g06460,At1g22370,At2g02990,At2g39030,At2g42530,At3g26580,At3g62550,At4g02370,At4g09020,At4g19120,At5g06690,At5g26340

400 - 600

185

1088

22229

7.0270

4.8945

0.121828404294242

At1g75190,At1g76590,At2g01520,At2g15970,At2g25930,At3g05880,At3g15630,At3g47860,At4g09020,At4g12280,At4g33700,At5g48250,At5g57630

600 - 800

185

1165

22229

11.8919

5.2409

0.000287070980858587

At1g29395,At1g48330,At1g51090,At1g69830,At1g75190,At1g76590,At2g17840,At2g34930,At2g47890,At3g10410,At3g28290,At3g46640,At3g50970,At3g59350,At4g12280,At4g23600,At4g35770,At5g07010,At5g25210,At5g50450,At5g52310,At5g62360

800 - 1000

185

1419

22229

9.7297

6.3836

0.0492800406452404

At1g26665,At2g16660,At2g21660,At2g22450,At2g28900,At2g38465,At2g42530,At2g42540,At3g22740,At3g46640,At3g50970,At3g53990,At4g11310,At4g30650,At5g03240,At5g15960,At5g47240,At5g52310

Table 2

Example of Frequency analysis option

Table 3 shows the distribution of different frequency states of DREB consensus sequence (RCCGAC) sequence among genes which are induced by draught stress( slide: drought_6h_shoot(1007966668#1029) , fold cutoff: 1.4).

CRE Frequency	Frequency of Occurrnce of CRE in class(Fc)	Number of CRE containing genes in class(Gc)	Number of genes in class(Tc)	Frequency of occurrence of CRE in reference set(Fr)	Number of CRE containing genes in reference set(Gr)	Number of genes in reference set(Tr)	Class Occurrence of CRE (Co=Fc/Tc)	Reference set occurrence of CRE (Ro=Fr/Tr)	Occurrence Ratio (=Co/Ro)	Percentage of CRE containing gene is class( (Gc/Tc) X 100 )	Percentage of CRE containing genes in reference set ( (Gr/Tr) X 100 )	Hypergeometic P-value	P-value estimated by bootstrap method	Genes which contain input CRE
1	51	51	185	4355	4355	22229	0.27568	0.19592	1.41	27.5676	19.5915	0.00525809532467544	0.01	At1g06460,At1g22370,At1g26665,At1g27630,At1g29395,At1g48330,At1g49720,At1g69830,At2g01520,At2g02990,At2g15880,At2g16660,At2g17840,At2g19450,At2g21660,At2g22450,At2g25930,At2g29630,At2g34930,At2g38465,At2g47890,At3g05880,At3g10410,At3g15630,At3g18080,At3g26580,At3g28290,At3g47860,At3g48360,At3g62550,At3g63160,At4g01130,At4g02370,At4g11310,At4g23600,At4g27440,At4g30650,At4g33700,At4g35770,At5g03240,At5g07010,At5g15960,At5g25210,At5g26340,At5g47240,At5g48250,At5g50450,At5g57110,At5g57630,At5g62360,At5g63810
2	14	14	185	712	712	22229	0.07568	0.03203	2.36	7.5676	3.2030	0.00261942026992649	<0.01	At1g51090,At1g75190,At1g76590,At2g15970,At2g28900,At2g39030,At2g42530,At2g42540,At3g22740,At3g53990,At3g59350,At4g12280,At4g19120,At5g06690
3	3	3	185	133	133	22229	0.01622	0.00598	2.71	1.6216	0.5983	0.0996678212609147	0.08	At3g46640,At3g50970,At4g09020
4	1	1	185	16	16	22229	0.00541	0.00072	7.51	0.5405	0.0720	0.125201681897977	0.13	At5g52310
5	0	0	185	5	5	22229	0	0.00022	-	0	0.0225	-	-	-

CRE Variant	Number of CRE containing genes in class(Gc)	Number of genes in class(Tc)	Number of CRE containing genes in reference set(Gr)	Number of genes in reference set(Tr)	Percentage of CRE containing gene is class( (Gc/Tc) X 100 )	Percentage of CRE containing genes in reference set ( (Gr/Tr) X 100 )	Hypergeometic P-value	Genes which contain input CRE
GCCGAC	32	185	2078	22229	17.2973	9.3481	0.000479464125515584	At1g51090,At1g76590,At2g02990,At2g15970,At2g16660,At2g25930,At2g28900,At2g39030,At2g42530,At2g42540,At3g18080,At3g46640,At3g47860,At3g50970,At3g53990,At3g59350,At3g63160,At4g01130,At4g02370,At4g09020,At4g11310,At4g12280,At4g19120,At4g30650,At4g33700,At5g06690,At5g07010,At5g25210,At5g52310,At5g57630,At5g62360,At5g63810
aCCGAC	51	185	3545	22229	27.5676	15.9476	3.98593423864847e-05	At1g06460,At1g22370,At1g26665,At1g27630,At1g29395,At1g48330,At1g49720,At1g69830,At1g75190,At1g76590,At2g01520,At2g15880,At2g15970,At2g17840,At2g19450,At2g21660,At2g22450,At2g29630,At2g34930,At2g38465,At2g39030,At2g42530,At2g42540,At2g47890,At3g05880,At3g10410,At3g15630,At3g22740,At3g26580,At3g28290,At3g46640,At3g48360,At3g50970,At3g53990,At3g59350,At3g62550,At4g09020,At4g12280,At4g19120,At4g23600,At4g27440,At4g35770,At5g03240,At5g06690,At5g15960,At5g26340,At5g47240,At5g48250,At5g50450,At5g52310,At5g57110
tCCGAC	26	185	3434	22229	14.0541	15.4483	0.730643779073995	At1g22370,At1g33970,At1g49720,At1g51940,At1g75190,At1g76590,At1g79440,At2g15890,At2g15960,At2g19450,At2g43550,At3g01310,At3g10410,At3g22740,At3g50700,At4g01130,At4g02370,At4g11310,At4g11600,At4g15210,At4g30690,At4g33980,At4g39090,At5g15960,At5g35735,At5g54960
cCCGAC	11	185	1693	22229	5.9459	7.6162	0.841664135891827	At1g07040,At1g53885,At2g01520,At2g28840,At2g43550,At3g28290,At3g47160,At4g09020,At4g30650,At4g33700,At5g03240
GaCGAC	32	185	4545	22229	17.2973	20.4463	0.878007079432752	At1g14250,At1g17665,At1g20010,At1g22740,At1g29395,At1g51090,At1g79440,At2g19450,At2g21660,At2g25930,At2g28840,At2g28900,At2g33830,At2g34930,At2g38465,At2g39330,At2g39900,At3g01310,At3g10410,At3g26580,At3g50700,At3g59350,At3g63160,At4g14230,At4g14270,At4g17470,At4g27440,At4g34950,At4g35770,At4g39260,At5g23660,At5g63810
GtCGAC	17	185	1601	22229	9.1892	7.2023	0.179949411621242	At1g10760,At2g15970,At2g17840,At2g21130,At2g33830,At2g34810,At2g38240,At2g43550,At2g45560,At3g47860,At4g15210,At4g34950,At4g39260,At5g05600,At5g23660,At5g61380,At5g62360
GgCGAC	11	185	1928	22229	5.9459	8.6734	0.933826693358881	At1g07040,At1g28050,At1g75190,At2g15830,At3g59930,At4g15210,At4g18240,At5g23240,At5g54960,At5g62360,At5g62720
GCaGAC	20	185	2742	22229	10.8108	12.3352	0.768224205053058	At1g02300,At1g06460,At1g22740,At1g22770,At1g70420,At1g77210,At2g25730,At2g43510,At3g52180,At3g59930,At3g63160,At4g11310,At4g12280,At4g14270,At4g15660,At4g30690,At5g05600,At5g48250,At5g57550,At5g62360
GCtGAC	24	185	2940	22229	12.9730	13.2260	0.573577158795241	At1g26665,At1g29395,At1g49720,At1g69830,At1g70420,At2g15830,At2g22450,At2g25730,At2g29630,At2g34930,At3g01310,At3g10410,At3g15450,At3g46640,At3g47340,At3g49620,At3g52180,At4g01130,At4g19390,At4g30660,At4g33980,At4g35770,At4g39260,At5g06870
GCgGAC	7	185	1066	22229	3.7838	4.7955	0.789104033261681	At1g06460,At1g22570,At2g34930,At3g07650,At5g11150,At5g54960,At5g67480
GCCaAC	35	185	4094	22229	18.9189	18.4174	0.459740187574122	At1g17665,At1g22370,At1g49720,At1g69830,At1g77210,At2g01520,At2g16660,At2g21130,At2g29630,At2g34930,At2g38240,At2g39330,At2g39900,At2g42540,At2g47890,At3g26740,At3g28290,At3g50970,At3g63160,At4g04330,At4g11310,At4g12280,At4g19390,At4g27440,At4g30650,At4g30660,At4g39090,At5g06690,At5g07010,At5g14550,At5g23660,At5g26340,At5g27280,At5g62720,At5g63810
GCCtAC	18	185	2092	22229	9.7297	9.4111	0.477495223930389	At1g26665,At1g52410,At1g68050,At2g28840,At2g33830,At2g38465,At2g42540,At3g07650,At3g59350,At4g12280,At4g14270,At4g17470,At4g35770,At5g14920,At5g26570,At5g48250,At5g61380,At5g63810
GCCcAC	28	185	2770	22229	15.1351	12.4612	0.159762078229407	At1g06460,At1g07040,At1g10760,At1g11210,At1g14250,At1g22740,At1g22770,At1g28330,At1g68050,At1g80480,At2g25930,At2g39900,At2g43550,At3g07650,At3g18080,At3g46640,At3g49620,At3g50970,At3g63160,At4g39260,At5g07010,At5g11150,At5g14550,At5g25210,At5g43440,At5g54960,At5g57110,At5g62360
GCCGtC	23	185	2376	22229	12.4324	10.6887	0.251716257316645	At1g28050,At1g28330,At1g33970,At1g69830,At1g79440,At2g25730,At2g28900,At2g43550,At3g29320,At3g49620,At3g53800,At3g62550,At4g01130,At4g02370,At4g12280,At4g30690,At4g35770,At4g39260,At5g25210,At5g42900,At5g59320,At5g64860,At5g67480
GCCGgC	4	185	750	22229	2.1622	3.3740	0.874512206479939	At1g20010,At3g49620,At4g15210,At4g15660
GCCGcC	22	185	1895	22229	11.8919	8.5249	0.0698733913297847	At1g02300,At1g22370,At1g33970,At2g25930,At2g42540,At2g43550,At2g45560,At3g26580,At3g46640,At3g49620,At3g50500,At3g50970,At3g59350,At3g59930,At3g63160,At4g11360,At4g11600,At4g12280,At4g19390,At4g26670,At5g35735,At5g54960
GCCGAa	18	185	2766	22229	9.7297	12.4432	0.894776958560156	At1g07050,At1g20010,At1g53885,At2g15830,At2g21660,At2g34810,At2g38240,At2g42540,At3g10410,At3g46640,At3g47160,At4g16146,At4g35770,At5g15960,At5g24470,At5g57110,At5g61380,At5g67480
GCCGAt	19	185	2455	22229	10.2703	11.0441	0.666599710422168	At1g06460,At1g14250,At1g20010,At2g25730,At2g34810,At2g34930,At2g43550,At2g47890,At3g62550,At4g02370,At4g14230,At4g16146,At4g35770,At5g15960,At5g43440,At5g54960,At5g57630,At5g62720,At5g67480
GCCGAg	14	185	2023	22229	7.5676	9.1007	0.801715143071705	At1g07050,At1g22370,At1g26665,At1g29395,At1g75190,At2g21660,At2g43550,At3g26580,At3g26740,At3g46970,At3g53800,At3g63160,At4g12280,At4g14270