Download SALSA MLPA probemix P018-F1 SHOX - MRC

MRC-Holland
Description version 18; 26-11-2012
®
MLPA
SALSA MLPA probemix P018-F1 SHOX
Lot F1-0411: As compared to version E1 (lot E1-1009), three new probes near the PAR1 boundary have
been included. One probe (GPR143) has been removed. The 88 and 96 nt control fragments have been
replaced (QDX2).
Léri-Weill Dyschondrosteosis (LWD) is a dominant skeletal disorder characterized by short stature and
distinct bone anomalies. Deletions of the SHOX gene and/or regulatory elements downstream of SHOX, are
detected in approximately 60% of LWD patients (Benito-Sanz et al. 2006; Huber et al. 2006; Fukami et al.
2006; Chen et al. 2009). The SHOX gene and the regulatory elements downstream of SHOX, are located
within the PAR1 region (pseudoautosomal region 1) which covers the 3000 kb of the X and Y chromosomes
next to the p-telomere.
This P018-F1 SHOX probemix contains probes for each exon of the human SHOX gene, as well as a probe
just before the SHOX promoter region. In addition, several probes are present detecting sequences in a
region downstream of SHOX which has been implicated in regulation of SHOX transcription. Furthermore,
several probes on the X chromosome are included in this probemix that can be used to characterise larger
deletions and to distinguish SHOX deletions from a Turner syndrome karyotype. Finally, ten autosomal
reference probes are included.
This SALSA® MLPA® probemix is designed to detect copy number changes of one or more sequences in the
aforementioned gene and chromosomal regions in a DNA sample. For sequences within the PAR regions and
the autosomal reference probes, heterozygous deletions of recognition sequences should give a 35-50%
reduced relative peak area of the amplification product of that probe. Deletions of a probe’s recognition
sequence on the X-chromosome, outside the PAR regions, will lead to a complete absence of the
corresponding probe amplification product in males, whereas female heterozygotes are recognisable by a
35-50% reduction in relative peak area. Note that a mutation or polymorphism in the sequence detected by
a probe can also cause a reduction in relative peak area, even when not located exactly on the ligation site!
In addition, some probe signals are more sensitive to sample purity and small changes in experimental
conditions. Therefore, deletions and duplications detected by MLPA should always be confirmed by other
methods. Not all deletions and duplications detected by MLPA will be pathogenic; users should always verify
the latest scientific literature when interpreting their findings. We have no information on what percentage
of defects in these genes is caused by deletions/duplications of complete exons. Finally, note that many
defects in the SHOX gene are expected to be small (point) mutations which will not be detected by this
SALSA® MLPA® test.
SALSA® MLPA® probemixes and reagents are sold by MRC-Holland for research purposes and to
demonstrate the possibilities of the MLPA technique. They are not CE/FDA certified for use in
diagnostic procedures. Purchase of the SALSA® MLPA® test probemixes and reagents includes a
limited license to use these products for research purposes.
The use of a SALSA® MLPA® probemix and reagents requires a thermocycler with heated lid and sequence
type electrophoresis equipment. Different fluorescent PCR primers are available. The MLPA technique has
been first described in Nucleic Acid Research 30, e57 (2002).
Related SALSA® MLPA® probemixes
P329 CRLF2-CSF2RA-IL3RA: contains probes for these PAR1 region genes. Deletions of the IL3RA and
CSF2RA genes, resulting in CRLF2 overexpression, are found in Acute Lymphoblastic Leukaemia samples.
P216 GHD: contains probes for GH1, PROP1, POU1F1, GHRHR, HESX1, LHX3 and LHX4.
P217 IGF1R: contains probes for IGF1R and IGFBP3.
P262 GHI: contains probes for IGF1, GHR, JAK2 and STAT5B.
P026 Sotos: contains probes for NSD1.
P384 Human Height: contains probes for HHIP, HMGA2, ADAMTSL3, NPPC, IHH.
More information
Website
E-mail
Mail
: www.mlpa.com
: [email protected] (information & technical questions); [email protected] (for orders)
: MRC-Holland bv; Willem Schoutenstraat 6, 1057 DN Amsterdam, the Netherlands
SALSA MLPA probemix P018 SHOX
Page 1 of 8
MRC-Holland
Description version 18; 26-11-2012
®
MLPA
References for SALSA® MLPA® probemix P018 SHOX
Chen, J. et al. (2009). Enhancer mutations of the SHOX gene as a frequent cause of short stature - the
essential role of a 250 kb downstream regulatory domain. J.Med.Genet. on line
Benito-Sanz S. et al. (2006). PAR1 deletions downstream of SHOX are the most frequent defect in a
Spanish cohort of Leri-Weill dyschondrosteosis (LWD) probands. Hum Mutat. 2006 Oct;27(10):1062.
Gatta V. et al. (2006). Identification and characterization of different SHOX gene deletions in patients with
Leri-Weill dyschondrosteosys by MLPA assay. J Hum Genet. 2006 Nov 8.
Fukami, M. et al. (2006). Transactivation function of an approximately 800-bp evolutionarily conserved
sequence at the SHOX 3' region: implication for the downstream enhancer. Am. J. Hum. Genet. 78: 167-170.
Huber et al. (2006). High incidence of SHOX anomalies in individuals with short stature. J Med Genet.
43(9):735-9.
Schiller et al. (2000) Phenotypic variation and genetic heterogeneity in Léri-Weill syndrome. Eur. J. Hum.
Genet. 8: 54-62
Exon numbering
The exon numbering from the Genbank mRNA reference sequences is used in this product description. This
exon numbering is different from the SHOX exon numbering in many articles. At least 2 alternatively spliced
SHOX mRNA forms are known, encoding proteins with different patterns of expression. The 2 mRNAs,
SHOXa and SHOXb, encode for proteins of 292 and 225 amino acids, respectively. Both transcripts have a
common 5' end. Transcript variant 1, as exemplified by the Genbank NM_000451.3 reference sequence,
contains exons 1-6. Transcript variant 2, as exemplified by reference sequence NM_006883.2, contains
exons 1-5 + 7. Exon 6 is called 6a and exon 7 is called exon 6b in most SHOX articles. Exon 7 contains the
last 42 nt of the variant 2 coding sequence. As this exon 7 sequence is entirely composed of two Alu
repeats, we designed a probe in a unique intron sequence located in front of exon 7.
Data analysis
The P018-F1 SHOX probemix contains 45 MLPA probes with amplification products between 130 and 490 nt.
In addition, it contains 10 control fragments generating an amplification product smaller than 120 nt: four
DNA Quantity fragments (Q-fragments) at 64-70-76-82 nt, three DNA denaturation control fragments (Dfragments) at 88-92-96 nt, one X-fragment at 100 nt and two Y-fragment at 105 nt and 118 nt. More
information on how to interpret observations on these control fragments can be found in the MLPA protocol.
Data generated by this probemix can be intra-normalized by dividing the peak area of each amplification
product by the total area of only the reference probes in the probemix (block normalization). Ratios can be
obtained by dividing the intra-normalized probe ratio in a sample by the average intra-normalized probe
ratio of all reference runs. This type of normalization assumes that no changes occur in the genomic regions
targeted by the reference probes. It is strongly recommended using reference and patient samples of the
same sex to minimize variation, as intersex comparison makes analysis more difficult. Sex determination can
also be done by visual examination of the electropherogram.
Data normalisation should be performed within one experiment. Only samples purified by the same method
should be compared. Confirmation of most exons deletions and amplifications can be done by e.g. Southern
blots or long range PCR.
Note that Coffalyser, the MLPA analysis tool developed at MRC-Holland, can be downloaded free of charge
from our website www.mlpa.com.
Many copy number alterations in healthy individuals are described in the database of genomic variants:
http://projects.tcag.ca/variation. For example, a duplication of a complete gene might not be pathogenic,
while a partial duplication or a deletion may result in disease. For some genes, certain in-frame deletions
may result in a very mild, or no disease. Copy number changes of reference probes are unlikely to be the
cause of the condition tested for. Users should always verify the latest scientific literature when interpreting
their findings.
This probemix was developed at MRC-Holland. Info/remarks/suggestions for improvement: [email protected].
SALSA MLPA probemix P018 SHOX
Page 2 of 8
MRC-Holland
®
MLPA
Description version 18; 26-11-2012
Table 1. SALSA MLPA P018-F1 SHOX probemix
Length
(nt)
64-70-76-82
88-92-96 *
100
105
118 *
SALSA MLPA probe
Chromosomal position
reference
Outside PAR1
SHOX region / PAR1
Q-fragments: DNA quantity; only visible with less than 100 ng sample DNA
D-fragments: Low signal of 88 or 96 nt fragment indicates incomplete denaturation
X-fragment: Specific for the X chromosome (AMOT gene)
Y-fragment: Specific for the Y chromosome (UTY gene)
Y-fragment S0135-L16766: Specific for the Y chromosome (ZFY gene)
130
Reference probe 00797-L00463
5q31
136
SHOX-area probe 05642-L05096
Xp22.33-PAR1
142
IL3RA probe 13597-L15055
Xp22.33-PAR1
148
SHOX-area probe 05648-L06218
Xp22.33-PAR1
154
SHOX-area probe 13821-L14642
Xp22.33-PAR1
160
Reference probe 04966-L04696
1p22
166
SHOX probe 01145-L00702
Exon 1 PAR1
172
SHOX-area probe 05643-L15705
Xp22.33-PAR1
178 ¥
SHOX-area probe 05649-L20176
Xp22.33-PAR1
185 ¥
SHOX-area probe 06293-L20177
Xp22.33-PAR1
191 ¥
Reference probe 06057-L05512
4p16
199 ¥
SHOX-area probe 13296-L20175
Xp22.33-PAR1
204
SHOX probe 01146-L06220
Exon 2 PAR1
211 ~
PPP2R3B probe 09333-L10292
Xp22.33-PAR1
219
Reference probe 03247-L02684
13q14
226 ¥
SHOX probe 09336-L20178
Intron 6 PAR1
231
SHOX probe 09337-L00911
Exon 6 (6a) PAR1
238 +
KAL1 probe 06402-L09795
Xp22.31
245
SHOX probe 01147-L00802
Exon 3 PAR1
254 **+
ARSF probe 16846-L20647
Xp22.33
261 ¥
Reference probe 00587-L20649
18q21
266 ¥
LOC159015 probe 01341-L20651
4 kb before SHOX-PAR1
274 +
FANCB probe 03906-L03066
Xp22.2
283 +
NLGN4X probe 05587-L04577
Xp22.31
290
SHOX-area probe 06291-L06222
Xp22.33-PAR1
300
SHOX probe 01148-L15501
Exon 4 PAR1
310 ~
ASMT probe 01153-L00712
Xp22.33-PAR1
318
SHOX-area probe 05645-L05099
Xp22.33-PAR1
328 **+
PRKX probe 16898-L19768
Xp22.33
337 ¥
SHOX probe 01149-L19676
Exon 5 PAR1
346
Reference probe 06560-L06118
1q32
VAMP7 (SYBL1) probe 01156-L00659
Xq28-PAR2
355 ∞
369 ¥
Reference probe 01838-L19680
16p13
377 ¥
SHOX-area probe 14697-L19677
Xp22.33-PAR1
386
CSF2RA probe 10251-L15502
Xp22.33-PAR1
392
SHOX probe 09338-L15503
Intron 6 PAR1
403 ¥
CRLF2 probe 13911-L19678
Xp22.33-PAR1
412
Reference probe 09793-L12593
15q21
420 +
AIFM1 (PDCD8) probe 00820-L15506
Xq26.1
432
SHOX-area probe 05646-L15507
Xp22.33-PAR1
442 ¥
SHOX-area probe 09335-L19679
Xp22.33-PAR1
453 **
ZBED1 probe 16858-L15742
Xp22.33-PAR1
463
SHOX-area probe 13297-L15510
Xp22.33-PAR1
474
Reference probe 09888-L10301
16p13
490 ±
Reference probe 12463-L13464
9q31
* Control fragments 88, 96 and 118 have been replaced in version F1 (from lot F1-0411 onwards).
** New in version F1 (from lot F1-0411 onwards).
¥ Changed in version F1 (from lot F1-0411 onwards). Change in length but no change in sequence detected.
+ X-chromosome, outside PAR region. Gives 50 % reduced signal in males as compared to females.
SALSA MLPA probemix P018 SHOX
Page 3 of 8
MRC-Holland
®
MLPA
Description version 18; 26-11-2012
~ This probe has been found to be duplicated in an apparently healthy individual in one of our quality tests.
∞ This probe has been found to be deleted in an apparently healthy individual in one of our quality tests.
± More variable. This probe has a high standard deviation
Table 2. SHOX probes arranged according to chromosomal location
Length SALSA MLPA
(nt)
probe
Gene
Exon
Ligation site
NM_000451.3
Partial sequence (24 nt
adjacent to ligation site)
Distance to
next probe
P-telomere / start PAR1 region
PPP2R3B gene
CGTCCGAGTTCC-ACTCGCGCTACA
SHOX region
4.7 kb before start
GCCTGGAACAGA-ACTTCCGCGGGG
LOC159015
SHOX gene
227 kb
273.0 kb
211 ~
09333-L10292
266
01341-L20651
166
204
245
300
337
231
01145-L00702
01146-L06220
01147-L00802
01148-L15501
01149-L19676
09337-L00911
SHOX stopcodon
1568-1570 (ex 6)
226
09336-L20178
SHOX intron 6
392
09338-L15503
SHOX intron 6
6.4 kb after exon 6
1.4 kb before exon 7
(only present in
NM_006883.2)
136
154
172
199
318 §
432 §
463 §
290 §
185 §
148
178
442
377
403
386
142
310 ~
453
05642-L05096
13821-L14642
05643-L15705
13296-L20175
05645-L05099
05646-L15507
13297-L15510
06291-L06222
06293-L20177
05648-L06218
05649-L20176
09335-L19679
14697-L19677
13911-L19678
10251-L15502
13597-L15055
01153-L00712
16858-L15742
Xp22.32-PAR1
Xp22.32-PAR1
Xp22.32-PAR1
Xp22.32-PAR1
Xp22.32-PAR1
Xp22.32-PAR1
Xp22.32-PAR1
Xp22.32-PAR1
Xp22.32-PAR1
Xp22.32-PAR1
Xp22.32-PAR1
Xp22.32-PAR1
Xp22.32-PAR1
CRLF2 gene
CSF2RA gene
IL3RA gene
ASMT gene
ZBED1 gene
GCAGCAGTGAAA-GTGAGCATTCCC
GATGGCTGATAA-TTACTCCGTATG
ACACCACCAGAGT-TACTTGAATCAA
GGAAAACCACGT-TCCTATCGATCC
TGTTCCCACCGT-AAAACTCACTCC
TGCATGTCTGCT-TTTTGAATGGCC
TACAGCAAATGA-TACGTATAAATT
CTTGAAAGGGCA-GGAACTCTAATT
TAATTGATGAGA-TGCAGAAGCCAG
TGGTGCTGAAAT-GAGGAAGCCCTG
TGAGGAGGTACC-TCAAAGCTAAAC
GAAATTCAGTTT-TAATAACACAGA
CTCTGGTGAGAT-GCCATCTAGAGA
GAATGCCAGCAA-ATACTCCAGGAC
GACAAGCCTTCT-GCTCTGTGAGTT
TGCACAGATAAG-TTTGTCGTCTTT
GACATCCCAGAA-GTGGTGTGGACG
TCGTCAAGAGCA-ACACGGAGCAGA
19.8
11.4
45.4
57.7
8.5
10.7
6.3
0.4
15.4
48.7
64.4
66.0
338.0
73.9
69.8
280.7
706.4
593.8
kb
kb
kb
kb
kb
kb
kb
kb
kb
kb
kb
kb
kb
kb
kb
kb
kb
kb
254 +
328 +
283 +
238 +
274 +
420 +
16846-L20647
16898-L19768
05587-L04577
06402-L09795
03906-L03066
00820-L15506
ARSF gene
PRKX gene
NLGN4X gene
KAL1 gene
FANCB gene
AIFM1 gene
End of PAR1 region
Xp22.33
CATCCATATAAT-TATGGGTTTGAC
Xp22.33
CGATTAGGAAAC-ATGAAGGTCAGT
Xp22.31
GACGGCTTGGGT-GATGCACGAAAT
Xp22.31
GTTTCCTGAAGC-GTGTGCCCACAA
Xp22.2
TCTCATCAGAAT-TCTCCCTATAAA
Xq26.1
TATTGGTCTTGT-GGACAGTAGTTT
Start of PAR2 region
536.9
2606.5
2311.9
6304.2
114321.4
25732.3
kb
kb
kb
kb
kb
kb
355∞‡
01156-L00659
VAMP7 gene (PAR2)
SHOX startcodon
692-694 (ex 2)
SHOX exon 1
SHOX exon 2
SHOX exon 3
SHOX exon 4
SHOX exon 5
SHOX exon 6 (6a)
99-100
920-921
1032-1034
1198-1199
1260-1261
1506-1507
Xq28
4.7 kb
TTTCTACTGCAA-ACAGAAATGGGA
ACCACGTAGACA-ATGACAAGGAGA
CGGGCAGACCAA-GCTGAAACAGAG
CAGAACCGGAGA-GCCAAGTGCCGC
ACAGCCAACCAC-CTAGACGCCTGC
AAGCAACAGCAA-GAATTCCAGCAT
6.7
3.6
6.2
0.2
3.5
6.4
TGGCTTCACGAG-TTCAGCCCATTG
6.4 kb
TCCCACATTCTT-GGAATCACAATG
56.8 kb
TGTGGGAAAAGT-GTTTCCATTCTG
Very close to
q-telomere
+ X-chromosome, outside PAR region. Gives halve the signal in males as compared to females.
~ This probe has been found to be duplicated in an apparently healthy individual in one of our quality tests.
∞ This probe has been found to be deleted in an apparently healthy individual in one of our quality tests.
‡ The VAMP7 probe at 355 nt is located very close to the q-telomere of X and Y in the PAR2 pseudoautosomal region.
§ These five probes are in the putative SHOX regulatory region (CNE7-9) described by Chen J. (2009) J.Med.Genet.
online. The 290 and 185 nt probes are within the putative regulatory region identified by Fukami, M. et al (2006) Am. J.
Hum. Genet. 78: 167-170.
SALSA MLPA probemix P018 SHOX
kb
kb
kb
kb
kb
kb
Page 4 of 8
MRC-Holland
Description version 18; 26-11-2012
®
MLPA
Note: Exon numbering is according to the NCBI NM_ reference sequences and might be different from the
literature! Several probes are only included to facilitate determination of the extent of a deletion /
duplication. Copy number alterations of certain probes may not be related to the condition being tested.
Complete probe sequences are available on request: [email protected]. Please notify us of any mistakes:
[email protected].
The following changes in the relative probe signals (as compared to the autosomal reference
probes indicated in table 1) are expected when MALE reference DNA is used:
1. An approx. 100% increase in signal of the chromosome X probes outside the PAR regions (marked + in
table 2) and a complete loss of the Y probe signals in female DNA. The signals of the SHOX probes, the
other probes in the PAR1 region and the VAMP7 (SYBL) probe in the PAR2 region at the q telomere
should remain unchanged as the Y chromosome also contains a copy of the PAR1 and PAR2 regions.
2. A 40-60% reduction in signal of one or more SHOX probes, in case of a (partial) deletion of the SHOX
gene. These deletions will in general cause Leri-Weill dyschondrosteosis.
3. A 40-60% reduction in signal of one or more probes downstream of SHOX, indicated with Xp22.32-PAR1
in table 2. Deletions in this region have been associated with Leri-Weill dyschondrosteosis (Benito-Sanz
et al (2005) Am. J. Hum. Genet. 77, 533-544). We included numerous probes in this region, as the exact
location of the SHOX regulatory region is not 100% certain. The most likely region is marked with § in
table 2. Please note that not all deletions detected by the Xp22.32-PAR1 probes will result in Leri—Weill
dyschondrosteosis.
4. A 40-60% reduction in signal of the SHOX probes and all other PAR1 probes and a complete loss of one
or more other Xp probes in case of deletions of Xp that extend outside PAR1.
5. A complete loss of the Y probe signals and a 40-60% reduction of the SHOX, the PAR1 and the PAR2
probes in Turner syndrome (X0).
6. An approx. 100 % increase in signal of the X probes outside the PAR regions and a 40-60% increase in
signal of the SHOX and the PAR1 and PAR2 probes in Klinefelter syndrome (XXY).
7. An approx. 200 % increase in signal of the X probes outside the PAR regions, the disappearance of the Y
probe signals and a 40-60% increase in signal of the SHOX and the PAR1 and PAR2 probes in Triple X
syndrome (XXX).
The following changes in the relative probe signals (as compared to the autosomal reference
probes indicated in table 1) are expected when FEMALE reference DNA is used:
1. A 40-60% decrease in signal of the chromosome X probes outside the PAR regions (marked + in table 2)
and the appearance of Y probe signals in male DNA. The signals of the SHOX probes, the other probes
in the PAR1 region and the VAMP7 (SYBL) probe in the PAR2 region at the q telomere should remain
unchanged as the Y chromosome also contains a copy of the PAR1 and PAR2 regions.
2. A 40-60% reduction in signal of one or more SHOX probes, in case of a (partial) deletion of the SHOX
gene. These deletions will in general cause Leri-Weill dyschondrosteosis.
3. A 40-60% reduction in signal of one or more probes downstream of SHOX, indicated with Xp22.32-PAR1
in table 2. Deletions in this region have been associated with Leri-Weill dyschondrosteosis (Benito-Sanz
et al (2005) Am. J. Hum. Genet. 77, 533-544). We included numerous probes in this region, as the exact
location of the SHOX regulatory region is not 100% certain. The most likely region is indicated in table 2.
Please note that not all deletions detected by the Xp22.32-PAR1 probes will result in Leri—Weill
dyschondrosteosis.
4. A 40-60% reduction in signal of the SHOX probes and all other PAR1 probes and a 40-60% decrease in
signal of one or more other Xp probes in case of deletions of Xp that extend outside the PAR1 region.
5. A 40-60% reduction of all chromosome X probes in Turner syndrome (X0).
6. The appearance of Y probe signals and a 40-60% increase in signal of the SHOX and the PAR1 and
PAR2 probes in Klinefelter syndrome (XXY).
7. A 40-60 % increase in signal of all chromosome X probes (all probes in table 2) in Triple X syndrome
(XXX).
SALSA MLPA probemix P018 SHOX
Page 5 of 8
MRC-Holland
®
MLPA
Description version 18; 26-11-2012
SALSA MLPA probemix P018-F1 SHOX sample pictures
40000
96.44
86.23
35000
266.26
230.57
91.16 105.55
30000
244.81
184.27
25000
217.99
198.22
177.37
171.49 190.37
134.01
128.19
158.06
151.73
164.55
146.12
141.10
20000
115.01
260.31
451.81
385.75
289.41
376.64 392.97
309.24
224.73
354.10
345.16
211.18
203.81
433.28
490.99
336.56
253.67
15000
474.87
441.95
326.25
237.65
420.39
281.43
274.12
10000
D ye S ign al
463.82
403.13
316.99
100.71
412.00
368.40
298.91
5000
0
50
100
150
200
250
300
350
400
450
500
Size (nt)
Figure 1. Capillary electrophoresis pattern from a sample of approximately 50 ng human male control DNA
analysed with SALSA MLPA probemix P018-F1 SHOX (lot F1-0411). The old MLPA buffer (replaced in
December 2012) was used. Vials with the old MLPA buffer have a white label.
50000
184.29
198.20
45000
217.99
40000
128.27
35000
158.10 171.53 190.40
151.79
177.40
134.11146.20
86.31
96.51
260.26
224.72
211.18
141.18
230.60
30000
91.20 105.60
266.23
25000
244.82
289.38
20000
309.21
100.75 115.08
164.57
298.89
203.82
15000
D ye S ign al
316.96
253.62
393.04
385.79
376.71
412.08
368.48
403.18
451.79
433.26
463.78
336.61
326.27
237.63
10000
354.13
345.16
420.39
281.40
274.10
441.99
490.92
474.80
5000
0
50
100
150
200
250
300
350
400
450
500
Size (nt)
Figure 2. Capillary electrophoresis pattern from a sample of approximately 50 ng human male control DNA
analysed with SALSA MLPA probemix P018-F1 SHOX (lot F1-0411). The new MLPA buffer (introduced in
December 2012) was used. Vials with the new MLPA buffer have a yellow label.
SALSA MLPA probemix P018 SHOX
Page 6 of 8
MRC-Holland
®
MLPA
Description version 18; 26-11-2012
85.97
35000
96.33
266.23
230.43
30000
253.65
100.61
184.25
91.07
244.66
25000
224.69
198.25
217.92
211.15
237.64
260.31
177.35
158.08 171.50
133.97
151.73
128.17
190.38
146.11
164.54
203.80
141.11
20000
309.19
289.34
326.21
451.88
420.46
385.87
376.67
354.13 368.39
345.20
281.35 298.81
274.07
316.98
393.00
412.00
433.43
463.84
403.19
491.03
336.47
474.92
15000
441.87
D ye S ign al
10000
5000
0
50
100
150
200
250
300
350
400
450
500
Size (nt)
Figure 3. Capillary electrophoresis pattern from a sample of approximately 50 ng human female control
DNA analysed with SALSA MLPA probemix P018-F1 SHOX (lot F1-0411). The old MLPA buffer (replaced in
December 2012) was used. Vials with the old MLPA buffer have a white label.
45000
184.27
198.26
40000
260.26
217.93
158.09
134.00
151.75 171.52
128.20
35000
211.17
100.66
96.36
25000
224.68
177.36
146.13
141.15
86.04
30000
190.38
230.45
266.19
91.11
253.60
244.66
20000
289.30
309.19
326.21
237.62
281.32 298.82
164.58
15000
203.82
274.03
316.97
393.03
385.87
420.46
376.70
354.14
412.02
368.43
433.38
345.19
403.18
451.85
463.82
491.04
336.50
441.88
474.89
D ye S ign al
10000
5000
0
50
100
150
200
250
300
350
400
450
500
Size (nt)
Figure 4. Capillary electrophoresis pattern from a sample of approximately 50 ng human female control
DNA analysed with SALSA MLPA probemix P018-F1 SHOX (lot F1-0411). The new MLPA buffer (introduced in
December 2012) was used. Vials with the new MLPA buffer have a yellow label.
SALSA MLPA probemix P018 SHOX
Page 7 of 8
MRC-Holland
Description version 18; 26-11-2012
®
MLPA
Implemented Changes –compared to the previous product description version(s).
Version 18 (48)
- Electropherogram pictures using the new MLPA buffer (introduced in December 2012) added.
Version 17 (48)
- Various minor textual changes.
- Mistake corrected in table 2, 337 nt probe 01149-L19676 (1261-1262 changed into 1260-1261).
- Data analysis method has been modified.
Version 16 (46)
- Additional related SALSA MLPA probemixes added on page 1.
- Minor textual changes on page 7 (Implemented Changes...).
Version 15 (45)
- This product description has been changed to incorporate a new version (lot number added, changes in
Table 1 and Table 2, new pictures included).
Version 14 (45)
- Minor textual change of page 5 (The following changes….).
Version 13 (45)
- This product description has been changed to incorporate a new version (lot number added, changes in
Table 1 and Table 2, new pictures included).
- Various minor textual changes on page 1; various minor layout changes. Tables have been numbered.
- Small changes of probe lengths in Table 1 & 2.
SALSA MLPA probemix P018 SHOX
Page 8 of 8