Table 1

Validation results of candidate genes.

Laboratory results


Gene

Alt. splicing hypothesis

RT-PCR

Sequencing

qRT-PCR AdCa

qRT-PCR SCC

Status


ADD3

Cassette exon

4/6

+

+

+

validated

ANTXR1

Alt. transcription start site

N/A

N/A

-

-

devalidated

CLSTN1

Cassette exon

4/6

+

+

-

validated

CTNND1

Cassette exon (5')

-

N/A

+

+

devalidated

Cassette exon (3')

2/6

+

-

-

devalidated

FN1

Cassette exon

5/5

+

+

+

validated

KIAA1217

Alt. transcription start site

N/A

N/A

+

+

validated

Intron retention

2/6

+

-

-

devalidated

MYH14

Cassette exon

-

+

-

-

devalidated

MYO18A

Cassette exon

4/6

+

+

-

validated

NCOR2

Alternative 5'-splice site

4/6

+

+

-

validated

NUMB

Cassette exon

5/6

+

+

+

validated

SLK

Cassette exon

5/6

+

+

+

validated

SYNE2

Cassette exon

6/6

+

+

-

validated

TPM1

Mutually exclusive exon (5')

N/A

+

+

+

validated

Mutually exclusive exon (3')

N/A

+

+

+

validated


A hypothesis of the mode of alternative splicing was formulated based on exon array expression profile and available transcript annotations. Events with alternative start of transcription were also considered, although they are not alternative splicing events by definition. RT-PCR was conducted using six samples; shown here is the number of samples that show an alternative splicing pattern that is in concordance with the exon array results. Exon-exon junctions in RT-PCR products were examined by sequencing. qRT-PCR was performed using transcript variant specific primer pairs for both AdCa and SCC. qRT-PCR results confirmed the exon array results (+) if they showed a significant (paired t-test p ≤ 0.05) or high (SI ≥ 2.0 or SI ≤ 2.0-1) difference of transcript variant expression that was in agreement with the exon array results. Differential splicing of a gene was considered as validated if at least one of the qRT-PCR results was positive. If available, RT-PCR had to confirm the exon array results in the majority of samples and also sequencing results had to be positive. Detailed result see additional file 11. N/A indicates that the experiment was not performed.

Langer et al. BMC Genomics 2010 11:676   doi:10.1186/1471-2164-11-676

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