Ohler et al. Supporting Dataset S2:
Detailed information on UNCOVER predictions verified by RT-PCR or EST evidence. 

This file refers to the experiments in figure 2a. Each prediction is referenced by a human Ensembl
ID set containing gene, transcript, and exon ID; this refers to the exon
on the 5' end of the analyzed intron. Sequences of the newly predicted exons
are given, along with its coordinates within the intron. Because
some of the verifications used primers internal to the skipped exons, we
have sometimes not inferred both 5' and 3' end experimentally.

Verified UNCOVER predictions with no prior EST evidence:
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Fig 2A lane i (brain)
eid-ENSE00000881911.1:gid-ENSG00000004866.5:tid-ENST00000265438.3
TCGCCAGTACCTACTGCAACATCTTTTCTCCCTACACAGCGACTCCAGCTTGGGAGGGCAGGGCCAGGGTTGTCACAGCTTCCCCTGTGGTGTCTGCCTGCCAAGCACAGCTCTGGAGTTAGCCCTGGGTGTGAG
pos 740-875, 5' end exact, 3' end exact
suppression of tumorigenicity 7
Primers: CCAAAGTCAGCAACAATATGC forward, TGAGGATTGAATTCCACAGC reverse

Fig 2A lane ii (brain)
eid-ENSE00000862512.1:gid-ENSG00000126217.3:tid-ENST00000261963.3
NTTCTCCTACCACGTNCTNACTCACATNGCCAGNACCGTGATGGAACGTACNAGGAGNNTNNCCACGTNCCATTTGGATGCTGCATTA
this intron contains two UNCOVER predictions corresponding to VEGA; 
however, the observed skipping event contains ~90bp non-matching insert
Guanine nucleotide exchange factor DBS
Primers: AAAGGAAAACAGACCCCCTAA forward, AATCTGCTCCTCTGCACTTG reverse

Fig 2A lane iii (brain)
eid-ENSE00001201432.1:gid-ENSG00000168781.5:tid-ENST00000335092.3
AGCTGCTGGATGACCAGCACCCTGTGGTCCGGTTGCTGCGCAGTTTTTCCTCTGACTGTACAGGGGGCCGGCCAGTCTCCTTGGATGCCACGCTGGCGCATCACCTGCACCAGTGCTCCTACCACCTGCGCCTCTTCCGGAACTGGCTGCGCTCAGGCCAGGATGACCCCGAGTGCCTCTACG
pos 3508-3690, 3' end exact, 5' end unknown
no description
Primers: GCCGTAAAACGATTTTCTGTG forward, CGCTGGCAGACTCTACTCAA reverse

Fig 2A lane iv (brain)
eid-ENSE00001146476.1:gid-ENSG00000168781.5:tid-ENST00000335092.3
GGGGTATACTGGTGGGGGCATCCGTNTTANGACTNCATGTNTACCGTNNAGGATGATCTACCGGNGNNGTGCCATGATTAATGGCCCAATGCCGTGGAACACGT
~90bp non-matching sequence insert; 
coupled with alternative 3' splicing of exon (shorter) at the same time; 
sequence missing in A3E: GTGTATACAGTGGGGCCAGATTATGCCCATGCTGAAG
no description
Primers: GAAAGACGGGGTCGTACATC forward, CTTTCCCCTCACTGTCTCGT reverse

Fig 2A lane v (HeLa)
eid-ENSE00001084095.4:gid-ENSG00000164402.2:tid-ENST00000331370.1
GCTGGATCTGGGTGTGTCTGGGAGGCTAGGCTGTGACAGTGAAGGAAGCTGTGTCCTCTTGCCAGCAT
pos 828-895, 3'end exact, 5' end predicted
Septin-like protein KIAA0202
Primers: CTAGGCTGTGACAGTGAAGGA forward, CAGAGGATGTTGAAGCTGAAG reverse


---------------------------------------------------------------

UNCOVER predictions with prior EST evidence but no annotation:

eid-ENSE00000663745.1:gid-ENSG00000103148.2:tid-ENST00000262313.2/6978-7010
CGAGCCATGTCCTCCAACCGATGAGCAATTGGTTGCAGG
pos 6977-7015; skipped exon, exact prediction
example EST ID: gi|15762590|gb|BI771012.1 
EST contains an extra 140 nt skipped exon matching a repetitive element
CGTHBA protein containing the major human alpha-globin regulatory element

eid-ENSE00001259630.4:gid-ENSG00000007392.5:tid-ENST00000293872.4/802-912
GTTCAGTGTATCTTTGCCTGCCTACATCAATCTGCAAGGGAGTTGCAGAAAGCCTCATGTTCATCGAGCC
pos 877-946 (3' end of exon wrong b/c of 1 nt deletion in human; 5' also wrong)
example EST ID: gi|18778192|gb|BM545792.1
LUC7-like isoform a; sarcoplasmic reticulum protein LUC7B1

eid-ENSE00001199798.1:gid-ENSG00000170558.1:tid-ENST00000269141.1/19210-19287
ATGGAGCACAGACTGTAGAAATGAAAAGCACAGAAGAAAGAGCTGTGTGAAAGAAATATTTAATCATAACATGAAAAACTCATAGTGGCAACTTTTCCATTATGTTAAATTTTCCTCATTTCTATGTGTTCTGAGTATGTGCTTAAAAAGCGAGGTTGTAATTTTCTATCTTTGTGCCCATCTCCCATCGGTTTTTGATTTTGTCTTTTTGAG
pos 19209-19419; ESTs suggest long alternative 3' end rather than skipping
example ID: gi|13732428|gb|BG210741.1|BG210741 
Neural-cadherin precursor

------------------------------------------------------------------------

UNCOVER predictions of alternative terminal exons -- only the 5' end has
been experimentally determined, the exon continues on the 3' end

Fig 2A lane vi 
eid-ENSE00001379673.1:gid-ENSG00000159140.5:tid-ENST00000321758.3
GCGTGTTCCCTGGAAAAGAGGGACGGATGAACCTGGAAG
TAAGTAAAAGACATTCTAGGTGTGTAGCATCAAGGCAGTTAATATCCAAGCATCAGCTTT
CTCTTTATACATCTACACTGCATGGCCTGCACCAAATAAGGAACTGAACCAGGGGTATGT
TTTTACCTCCACAGCTGCCTCCTTCCATCAGAGCACCTTGATGAACTTAATGTCTAGTCA
CACGTCATTGGCATGTTTTCTCCCCAGCATTTAATT
SON protein (SON3)
Primers: CTCTATTCCTGGCCAGTTCA forward, ATCAAGGTGCTCTGATGGAA reverse

Fig 2A lane vii
eid-ENSE00001046164.1:gid-ENSG00000067369.1:tid-ENST00000263801.1
TGCTGGGAGGTAACAATCATGCCGTCT
GGAGTCTCCTGCTACCTATGCCTGATGCTGGCAGATGTGGGTTGGCTGAGGGCTCATGCA
GCATCCTTTGTCCAGATGCAGAAGTGGAACTGAATCCTGTTGC
Tumor suppressor p53-binding protein 1 (TP53BP1)
Primers: CCCATTTCATTTCACTTTGC forward, GCAACAGGATTCAGTTCCAC reverse

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Sequence conservation of newly verified skipped exons in other vertebrtates:

lane  ENSE      rat   dog   chicken

i      881911   Y     Y     Y       
ii     862512   N     N     N  (*)     
iii   1201432   Y     Y     Y       
iv    1146476   N     N     N  (*)     
v     1084095   Y     N     N

vi    1379673   Y     Y     N     
vii   1046164   Y     Y     N       


(*) amplified human sequence does not match the current genome assembly

-------------------------------------------------------------------

Random controls:

Fig 2A lane i
eid-ENSE00001321652.4:gid-ENSG00000161980.2:tid-ENST00000293860.2
Primers: AGACCATGCTGCTGTTCTG forward, ACCAAGCACATCATCCACTT reverse

Fig 2A lane ii
eid-ENSE00000868377.2:gid-ENSG00000102125.4:tid-ENST00000218246.3
Primers: GCTTCACCAAGGAGCTACAC forward, GGTTGAGCTTCTCCAAAATG reverse

Fig 2A lane iii
eid-ENSE00001239587.1:gid-ENSG00000100220.2:tid-ENST00000216038.2
Primers: CCCCAGAGGGTCAAGACTAT forward, CACTTTGGCAATGTTGTGAG reverse

Fig 2A lane iv
eid-ENSE00001307891.1:gid-ENSG00000185721.1:tid-ENST00000331457.1
Primers: GCCTCACTGTGTACCCATCT forward, TGTGGTCCTGGAGTAAGGAA reverse