Service items

　

Introduction of service types in Affymetrix Gene Expression Service Lab

 

Currently, there are 5 major service items provided by Affymetrix Gene Expression Service Lab (AGESL) as listed below.  Users are welcome to contact AGESL for service of items not listed.

To ensure continuous support of Affymetrix Gene Expression Service Lab (AGESL) from Academia Sinica, we urge you to acknowledge AGESL in your publication with data generated in our service lab.  Please use the following statement in your Acknowledgement section:

Acknowledgement

Affymetrix GeneChip assay were performed by the Affymetrix Gene Expression Service Lab (http://ipmb.sinica.edu.tw/affy/), supported by Academia Sinica.

For the user who has used AGESL and acknowledged AGESL in your publication, please fill out the application form to earn your 10% discount in the next ordering.

 

1. Eukaryotic 3’- Expression analysis

Provide service with 27 Affymetrix 3’- expression arrays (2009/5/8).  This service contains cDNA synthesis, target amplification and labeling, fragmentation, hybridization and image acquisition.  The starting material is high quality RNA (total RNA is recommended).

The RNA samples to be submitted should be resuspended in nuclease-free water and pass the 1st QC criteria.  Please label the “ID” on the top of 1.5 ml vial for each sample submitted.

Table1. 1st QC criteria of Eukaryotic 3’- Expression analysis samples

No

QC criteria

Threshold

1

Sample type

Total RNA or mRNA (total RNA is recommended)

2

Ratio of 260/230

1.8-2.5

3

Ratio of 260/280

1.9-2.5

4

Concentration (ug/ul)

0.1-2.0

5

RNA amounts (ug)

3-10

6

RNA volume (ul)

5-30

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2. Prokaryotic 3’- Expression analysis

Provide service with Affymetrix 3’- expression arrays.  This service contains DNA labeling, hybridization and image acquisition.  The starting material is DNase digested DNA (size of 50-200 bp is recommended).  Users have to decide the amount of DNA used for hybridization.

The DNA samples submited should be in nuclease-free water and pass the 1st QC criteria.  Please label the “ID” on the top of 1.5 ml vial for each sample submitted.

Table2. 1st QC criteria of Prokaryotic 3’- Expression analysis samples

No

QC criteria

Threshold

1

Sample type

DNase digested DNA

2

Ratio of 260/230

1.8-2.5

3

Ratio of 260/280

1.7-2.5

4

DNA concentration (ug/ul)

0.1-0.4

5

Fragmentized DNA amount (ug)

2-8

6

Fragmentized DNA size (bp)

50-200

7

DNA amount for hybridization (ug)

1-7

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3. Whole-Transcript Expression with Exon array

Provide service with Affymetrix Exon arrays.  This service contains rRNA depletion, cDNA synthesis, target amplification, fragmentation, labeling, hybridization and image acquisition.  The starting material is high quality RNA (total RNA is recommended).

The RNA samples submitted should be in nuclease-free water and pass the 1st QC criteria.  Please label the “ID” on the top of 1.5 ml vial for each submitted sample.

Table3. 1st QC criteria of Whole-Transcript Expression with Exon array

No

QC criteria

Threshold

1

Sample type

Total RNA or mRNA (total RNA is recommended)

2

Ratio of 260/230

1.8-2.5

3

Ratio of 260/280

1.9-2.5

4

Concentration (ug/ul)

0.1-2.0

5

RNA amounts (ug)

3-10

6

RNA volume (ul)

5-30

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4. Transcript Mapping with Tiling array (Single array)*

Provide service with Affymetrix tiling arrays for the discovery of novel transcripts and other whole-genome experiments.  This service contains cDNA synthesis, target amplification, fragmentation, labeling, hybridization and image acquisition.  The starting material is high quality RNA (total RNA is recommended).

The RNA samples submited should be in nuclease-free water and pass the 1st QC criteria.  Please label the “ID” on the top of 1.5 ml vial for each sample submitted.

Table4. 1st QC criteria of Transcript Mapping with Tiling array

No

QC criteria

Threshold

1

Sample type

Total RNA or mRNA (total RNA is recommended)

2

Ratio of 260/230

1.8-2.5

3

Ratio of 260/280

1.9-2.5

4

Concentration (ug/ul)

1.5-2.0

5

RNA amounts (ug)

9-18

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5. ChIP-on-chip with Tiling array (Single array)*

Provide service with Affymetrix tiling arrays for mapping sites of protein/DNA interaction in chromatin immunoprecipitation (ChIP) experiments, and other whole-genome experiments.  This service contains target fragmentation, labeling, hybridization and image acquisition.  The starting material is dUTP incorporated DNA.

The dUTP incorporated DNA submitted should be in nuclease-free water and pass the 1st QC criteria.  Please label the “ID” on the top of 1.5 ml vial for each submitted sample.

Table5. 1st QC criteria of ChIP-on-chip with Tiling array (Single array)*

No

QC criteria

Threshold

1

Sample type

dUTP incorporated DNA

2

Ratio of 260/230

1.8-2.5

3

Ratio of 260/280

1.7-2.5

4

dUTP incorporated DNA concentration (ug/ul)

0.2-0.5

5

dUTP incorporated DNA amounts (ug)

10-20

6

dUTP incorporated DNA size (bp)

200-2000

 

*If you need the service with multiple array set, please contact with affy@gate.sinica.edu.tw

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6. miRNA expression analysis

Provide service with Affymetrix miRNA arrays. This service contains miRNA labeling, hybridization and image acquisition. The starting material is high quality RNA (total RNA is recommended).

The RNA samples to be submitted should be resuspended in nuclease-free water and pass the 1st QC criteria.  Please label the “ID” on the top of 1.5 ml vial for each sample submitted.

Table1. 1st QC criteria of miRNA expression analysis samples

No

QC criteria

Threshold

1

Sample type

Total RNA or mRNA (total RNA is recommended)

2

Ratio of 260/230

1.8-2.5

3

Ratio of 260/280

1.9-2.5

4

Concentration (ug/ul)

0.1-2.0

5

RNA amounts (ug)

1.0-5.0

6

RNA volume (ul)

5-30

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7. Array-CGH expression analysis

Provide service with Affymetrix arrays for the detect deletions or duplications in chromosomes. This service contains DNA labeling, hybridization and image acquisition. The starting material is genomic DNA.

The DNA samples to be submitted should be resuspended in nuclease-free water and pass the 1st QC criteria.  Please label the “ID” on the top of 1.5 ml vial for each sample submitted.

Table1. 1st QC criteria of DNA expression analysis samples

No

QC criteria

Threshold

1

Sample type

genomic DNA

2

Ratio of 260/230

1.8-2.5

3

Ratio of 260/280

1.7-2.5

4

DNA concentration (ug/ul)

0.1-1.5

5

DNA amounts (ug)

3-10

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The genomic DNA contamination of samples is rarely to be detected by lab 1st QC metrics.  It is recommended for user to confirm it before submission (Please go to “FAQ” for more information).

Criteria for RNA integrity:

·         If your samples are total RNA, we will use a Bioanalyzer 2100 for analysis control. The criteria of Bioanalyzer 2100 results are:

28S peak > 18S peak

28S/18S ratio ≥ 1.4

18S+28S total area percentage ≥ 40

·         If your samples are mRNA, we will conduct the quality control procedure after finishing in vitro transcription.

　

Suggested RNA sample preparation methodology

species or tissue

RNA isolation

Purification of total RNA (2nd)

total RNA

polyA-RNA

Yeast

Hot phenol protocol. (Schmitt, et al.,1990, Nucl Acids Res, 18:3091-3092.)

Qiagen's Oligotex mRNA Kit

Qiagen's RNeasy Mini Kit

Arabidopsis

TRIzol Reagent (Invitrogen Life Tech.)

Qiagen's Oligotex mRNA Kit

Qiagen's RNeasy Mini Kit

Mammalian cells (cultured cells and lymphocytes)

Qiagen's RNeasy Mini Kit

Qiagen's Oligotex Direct mRNA Kit (directly from cell) or Qiagen's Oligotex mRNA Kit (from total RNA)

N/A

Mammalian tissue

TRIzol Reagent from Invitrogen Life Tech.

poly-A mRNA isolation commercial kit (from total RNA)

Qiagen's RNeasy Mini Kit

Albino or  polysacchoride rich in plant

Pine tree method

N/A

N/A

(Adapted from GeneChip® Expression Analysis Technical Manual, Affymetrix company)

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