Leiden Muscular Dystrophy pages©

SGCD primers for mutation screening

(last modified March 7, 2004)


Contents



DNA

DNA-primers

Amplified

Length

Reference

Primer forward /
primer reverse

Name

exon 2 227 Tsubata cctgccttctggaagtaatc /
aaaatgaccatgagcagggc
D-Sarc2F/R
exon 3 246 Tsubata tgcttctctcttgcctcgtt /
gctaaacaaacctagatggt
D-Sarc3F/R
  Nigro cagcggtttaatgtgagtgcttct /
ccggaagaaagctaaacaaacctaga
2F/2R

187

Duggan

TGCCTCAGGAGCAGTACACT /
 AATTGTGAAGTTCATGACTTTG

deltaE2F/R

exon 4 208 Tsubata ttacagcctgaggtgttttg /
gcaacaataatgcctccttc
D-Sarc4F/R
  Nigro tcatatcttccctgttatctctgtg /
caataatgcctccttctctcagcag
3F/3R
exon 5 222 Tsubata ccccttggagagttgtaatg /
tattctgagtgcctcgcatg
D-Sarc5F/R
  Nigro tcatgtctttctcttattttcttattg /
ggttcatatggtgtttcctttgacc
4F/4R
exon 6 245 Tsubata gatgagactaatggtgtttt /
aaaatgtacacaGTagcatc*
D-Sarc6F/R
  Nigro gcgatgagactaatggtgttttctc /
tgggaggacatatcctgttcataaca
5F/5R
exon 7 188 Tsubata caggtgactccagtatctcc /
tggccagttgcacagagcaa
D-Sarc7F/R
  Nigro caggtgactccagtatctccaatc /
gcacagagcaaggcaatagccatt
6F/6R
exon 8 199 Tsubata aaaagggatctttattgacg /
tgtagctctttgaaCttctgt*
D-Sarc8F/R
  Nigro ccttgagcatgaacttccttttgt  /
tggcctgttgaagctgtagctct
7F/7R
exon 8b 174 Tsubata aagctggcaatatggaagcc /
ggctccttttgttgatacac
D-Sarc9pF/R
exon 9 243 Tsubata ctgaccaatgctttccttcc /
atgctgccaacaatgtccac
D-Sarc9mF/R
    Nigro tctgaatgtcaagagaagagacgacag /
cagccagtgtgtaaagccaaaaa
8F/8R

Legend: (primer sequences obtained from Nigro, with the assistence of Eloisa de Sa Moreira)
*primer sequences listed have genomic targets but differ from those published (Tsubata, D-Sarc6R has GT in stead of TG, D-Sarc8F/R has a C inserted).
Exonic sequences are in upper case, intronic and gene flanking sequences in lower case and added primer tails in italics. Amplified: region amplified. Numbering of exons is according to Tsubata et al. Length: length of PCR-product in basepairs. Reference: publication describing the primer(s). Forward primer: sequence of forward primer. Reverse primer: sequence of reverse primer. Name: name of the primers.
PCR-conditions (Nigro primers); initial denaturation 30 sec at 99oC, 30 cycles with denaturation 15 sec at 96oC and 3 min annealing-extension at 61oC.


SSCA

ExonForward primer /
reverse primer
Size (bp)Amplification
2cctgccttctggaagtaatc / aaaatgaccatgagcagggc22965oC to 55oC (-2oC/2 cycles, 1 min); +30 cycles, 55oC, 1 min
3tgcttctctcttgcctcgtt / gctaaacaaacctagatggt24370oC to 60oC (-1oC/2 cycles, 1 min); + 30 cycles, 60oC, 1 min
4ttacagcctgaggtgttttg / gcaacaataatgcctccttc20870oC to 60oC (-1oC/2 cycles, 1 min); + 30 cycles, 60oC, 1 min
5ccccttggagagttgtaatg / tattctgagtgcctcgcatg21865oC to 55oC (-1oC/2 cycles, 1 min); + 30 cycles, 55oC, 1 min
6gatgagactaatggtgtttt / aaaatgtacacaGTagcatc *24465oC to 55oC (-2oC/2 cycles, 1 min); + 30 cycles, 55oC, 1 min
7caggtgactccagtatctcc / tggccagttgcacagagcaa18865oC to 55oC (-2oC/2 cycles, 1 min); + 30 cycles, 55oC, 1 min
8aaaagggatctttattgacg / tgtagctctttgaaCttctgt **19660oC to 50oC (-2oC/2 cycles, 1 min); + 30 cycles, 50oC, 1 min
9actgaccaatgctttccttcc / ATGCTGCCAACAATGTCCAC23960oC to 50oC (-2oC/2 cycles, 1 min); + 30 cycles, 50oC, 1 min
9bAAGCTGGCAATATGGAAGCC /
GGCTCCTTTTGTTGATACAC
17360oC to 50oC (-2oC/2 cycles, 1 min); + 30 cycles, 50oC, 1 min

Legend:
Primers for PCR and SSCA of the SGCD gene  (Sylvius et al., 2003). The primer sequences published (*aaaatgtacacaTGagcatc and **tgtagctctttgaa_ttctgt) differ from the human genome sequence as shown here.


RNA-primers

RT-PCR

Amplified

Reference

Length

Forward primer

Reverse primer

Name

-26-278

Duggan

304

GTGTTGAGTGAAGGGACCAG

TCTTTGGCGTAGAGAGGTTG

delta1F/R

192-486

Duggan

294

TGATGGAATGGGAAACCTGA

TTCAGCTCCTACTACCACTTCA

delta2F/R

436-748

Duggan

312

GGAAAATTGCTCTTCTCTGCA

ATCCATGAGGCAGTCTAGGT

delta3F/R

643-938

Duggan

295

GGCAATATGGAAGCCACCTG

GCAGGCAGTGTGTAAAGCCA

delta4F/R

Legend:
Exonic sequences are in upper case, intronic and gene flanking sequences in lower case and added primer tails in italics. Amplified: region amplified. Numbering of exons is according to Tsubata et al. Length: length of PCR-product in basepairs. Reference: publication describing the primer(s). Forward primer: sequence of forward primer. Reverse primer: sequence of reverse primer. Name: name of the primers.



| Top of page | LMDp homepage |
| NMD mutation detection | Muscular dystrophies |
| Remarks / information | Copyright©, liability |