Discrepant cytogenetic, MLPA and array CGH results on a der(X)(pter->q27.2::p22.31->pter): the

advantages of an integrated approach.

• M N Collinson, S Huang, JN Macpherson, VK Maloney, AC Skinner, A Karcanias, NA Affara, NS Thomas

30 year old female with premature ovarian failure (POF)

cytogenetic analysis molecular genetic testing for a FRAXA premutation

46,XX only one allele found?45,X or 46,XY

MLPA with the Rett syndrome kit

MRC-Holland kit PO15C

LIST OF PROBES USED FOR FISH TO CONFIRM DELETIONCentromere

RP1-142F18 Normal (Xq27.2)

RP1-315J21 Deleted (Xq27.3)

RP11-79A21 Deleted (Xq27.3)

RP11-37P24 Deleted (fragile X locus)

RP11-161L9 Deleted (fragile X locus)

RP11-64L19 Deleted (Xq28)

RP11-533H4 Deleted (Xq28)

RP11-76K17 Deleted (Xq28)

RP11-402H20 Deleted (Xq28)

Telomere

FISH with a probe for the FRAXA gene

RP11-161L9X centromere (control)

Analyst Slide

1

Slide

2

Slide

3

Slide

4

Slide

5

Slide

6

1 X

2 X

3 X

Blind cytogenetic analysis – 3 control slides and 3 slides from POF patient scored by 3 independent analysts

Partial karyotype of the X chromosomes(abnormal X on the right)

MLPA with the subtelomere probe kit

MRC-Holland kit PO36B

CY29 (Xp subtelomere)

c8.1/2 (Xq subtelomere)

FISH with the Xp and Xq subtelomere probes

Agilent 244K oligoarray

Array-CGH

RP11-946P8

DXZ2 (X centromere control)

FISH with SHOX probe

MLPA with SHOX kit PO18BDuplication of SHOX

POF case

FEMALE CONTROL MALE CONTROL

Array CGH with chip specifically designed for sex chromosome abnormalities

NORMAL CASE

FEMALE CONTROL MALE CONTROL

Array CGH with chip specifically designed for sex chromosome abnormalities

POF CASE

L.M. Russell et al, Cytogenet Genome Res 116:181-185 (2007)

Original cytogenetic analysis – 1/30 metaphases 45,X

Blind FISH study with X centromere probe (DXZ2)

Metaphases

Interphase cells

POF case Control

1/30 45,X (3.3%) 1/30 45,X (3.3%)

8/100 45,X (8%) 6/100 45,X (6%)

Summary• An integrated approach was required to establish the

true nature of the X chromosome abnormality

• Highlights the fact that one should be cautious of any shifts from the baseline when analysing array CGH as it may mask significant changes in log ratios

• We still do not have a satisfactory explanation of our results other than mosaicism

• Deletions of this region of Xq have been reported in association with POF