1. PRENATAL DIAGNOSIS
Dr.Ankur Puri
25/10/12
1

2. INTRODUCTION
The frequency of inherited malformations as well as
genetic disorders in newborns account for around 3-
5%.These frequency is much higher in early stages of
pregnancy, because serious malformations & genetic
disorders usually lead to spontaneous abortion.
Thereafter, taking into account the severity of the
disorders the decision should be taken in regard of
subsequent course of the pregnancy , possibilities of
treatment, parent's acceptation of a handicapped child
but also, in some cases the possibility of termination of
the pregnancy according to MTP act 1971.
2

3. • Definition:
‘Prenatal diagnosis is defined as the detection of
abnormalities in the fetus, before birth’
3

4. The purpose of prenatal diagnosis is not simply to detect
abnormalities in fetal life and allow termination.It rather
have following goals :
 Provide a range of informed choice to the couples at
risk of having a child with abnormality.
 Provide reassurance & remove anxiety, especially
among high risk groups.
 Allow couples at high risk to know that the presence or
absence of the disorder can be confirmed by testing.
 Allow the couples the option of appropriate
management (
psychological, pregnancy/delivery, postnatal)
 To enable prenatal treatment of the fetus. 4

5. Some Disorders for which PRENATAL DIAGNOSIS is
available:
1. Congenital malformations
2. Chromosomal disorders
3. Non genetic Fetal disorders
*Fetal infections, Immune hydrops, DM,Fetal effects of maternal drugs e.g valproic
acid
4. Single gene disorders
-Multiple malformation synd
*Holt oram, Craniosynostosis, Orofacial digital synd
-Hematological disorders
*Thalassemias, Hemoglobinopathies, Hemophilia
-Metabolic Disorders
*Tay sach, Wilson, MPS, CAH.
-Neuromuscular disorders
5
*Huntington chorea, Myotonic dystrophy, DMD, Fragile X

6. -Renal Disoders
*AD/AR polycystic kidney disease
-Connective tissue dis / Skeletal dysplasia
* Osteogenesis imperfecta, Ehlers Danlos, Achondroplasia, Marfan.
-Skin disorders
*Epidermolysis bullosa, Ichthyosis, Ectodermal dysplasia
6

7. INDICATIONS OF PRENATAL DIAGNOSIS
1. Advanced maternal age.
2. Previous child with a chromosomal abnormality.
3. Family history of a chromosomal abnormality.
4. Family history of a single gene disorder.
5. Family history Neural Tube Defect.
6. Family history of other congenital structural
abnormality.
7. Abnormalities identified in pregnancy.
8. Other risk factors(consanguinity,poor obs.
History,maternal history)
7

8. 1. Advanced maternal age
• It is the common indication for prenatal diagnosis.
• As a woman’s age increases, so does the risk for
chromosome aneuploidy in the fetus.
• Most centers offer Amniocentesis or CVS to a women aged
>35yrs,although no standard criterion exists at what age
women should be investigated.
8

9. 2. Previous child with a chromosomal
abnormality
• Previous child with Down’s So due to non dysjunction or
unbalanced translocation will give a risk in subsequent
pregnancy as, of mother’s age related risk is plus 5%.
• If one of the parents have balanced chromosomal
rearrangement (translocation, inversion) causing a serious
problem for a previous child due to unbalanced
rearrangement, then recurrence risk is between 1-2% & 15-
20 %.This risk will depend on nature of rearrangement &
nature of segment involved.
9

10. 3. Family history of a chromosomal
abnormality
• Usually no increase in risk compared to general population
since most chromosomal disorders will arise as a result of
disjunction than familial rearrangement.
• A history of Down’s So.
• However each situation should be confirmed by nature of
chromosome abnormality in affected individual or urgent
chromosomal analysis from blood of related parents if
normal,no invasive tests.
10

11. 4. Family history of a single gene
disorder.
• A previous affected child
• Affection of one of the parents
• Postive family history.
• Have a 25-50 % recurrence and prenatal diagnosis should
be offered as many can be diagnosed by DNA analysis and
Biochemical testing(achondroplasia, huntington disease,
neurofibromatosis)
11

12. 5. Family history Neural Tube Defect.
• In the 1st & 2nd degree relatives the risk should be
determined
• High risks were diagnosed by Amniocentesis & AFP
assessment.
• Ultrasound with MSAFP is method of choice now a days
12

13. 6. Family history of other congenital
structural abnormality.
• Evaluation of family pedigree
• Calculation of the risk.
• If increased risk-detailed ultrasound can be offered
between 16-18 weeks of pregnancy, it will detect most
serious defects (cranial, cardiac, renal & limb deformity)
13

14. 7. Abnormalities identified in
pregnancy
• Uncertainty of maternal serum screening & fetal anomaly
scanning can make invasive procedure for the diagnosis
more necessary.
• Poor fetal growth can be indication for prenatal
chromosome analysis as well as for confirmation of a
serious & non viable abnormality.
14

15. 8. Other risk factors(consaguinity,poor
obs. History,maternal history)
• Parental consanguinity leading to hereditary disorder or
congenital anomalies(offer a detailed USG)
• Poor obst history as recurrent miscarriage or still birth
indicating high risk in future pregnancy(offer USG of fetus &
chromosomal analysis of parents)
• Maternal illness as poorly controlled DM or maternal
epilepsy treated with some drugs such as sodium
valproate(offer a detailed USG)
15

16. METHODS OF PRENATAL DIAGNOSIS
NON INVASIVE INVASIVE TECHNIQUES
TECHNIQUES Fetal visualization
Fetal visualization Fetal tissue sampling
Maternal serum screening Cytogenetics
Separation of fetal cells Molecular genetics
from the mother's blood
16

17. NON INVASIVE TECHNIQUES
FETAL VISUALISATION
1. ULTRASONOGRAPHY
2. FETAL ECHOCARDIOGRAPHY
3. MAGNETIC RESONANCE IMAGING (MRI)
17

18. FETAL VISUALISATION
1. ULTRASONOGRAPHY :
-It is a noninvasive procedure for imaging fetal anatomy & is
harmless to both the fetus and the mother.
-The developing embryo can first be visualized at about 6
weeks gestation. Recognition of the major internal organs &
extremities to determine if any are abnormal can best be
accomplished between 16 to 20 weeks gestation.
- Thus USG is used in the 2nd trimester to identify major fetal
structural anomalies & fetal anatomical markers.
-Ultrasound also is used to guide invasive sampling, such as
amniocentesis, CVS, cordocentesis, & various fetal biopsies
18

19. US markers of fetal congenital abnormalities or genetic syndromes
found in first trimester scanning [at 11-13weeks' gestation]
19

20. 2D US
20

21. 2D US
21

22. 2D US
22

23. 2D US
23

24. 2D US
24

25. 3D & 4D US
• In recent years three-dimensional ultrasound (3D) &
four-dimensional ultrasound (4D) have started to
play an increasing role in prenatal diagnosis. They
can be applied in assessing facial features, central
nervous system abnormalities and skeletal defects
25

26. ‹#›

27. Ultrasonography cont…
• Although an ultrasound examination can be quite
useful to determine the size & position of the
fetus, the size & position of the placenta, the
amount of amniotic fluid, & the appearance of fetal
anatomy, there are limitations to this procedure as
findings are based upon views of the fetus, the
estimated gestational age, sonographer
experience, & the degree of anomaly severity.
27

28. FETAL VISUALISATION
2. FETAL ECHOCARDIOGRAPHY
-Fetal echocardiography is capable of diagnosing most significant
congenital heart lesions as early as 17-19 wk of gestation.
-When this technique is used with duplex or color flow Doppler,
it can identify a number of major structural cardiac defects &
rhythm.
-Fetal echocardiography is recommended in cases where cardiac
defects are suspected.
28

29. FETAL ECHOCARDIOGRAPHY : cont…
29

30. FETAL VISUALISATION
3. MAGNETIC RESONANCE IMAGING (MRI)
• MRI is used in combination with ultrasound, usually
at or after 18 weeks‘ gestation. MRI provides a tool
for examination of fetuses with large or complex
anomalies, and visualization of the abnormality in
relation to the entire body of the fetus. Apparently
MRI is a risk-free method
30

31. MAGNETIC RESONANCE IMAGING (MRI)
31

32. MATERNAL SERUM SCREENING
 Maternal serum screening is used to identify women at
increased risk of having a child with trisomies 18 or 21 or an
open neural tube defect (NTD), while posing no risk to
the pregnancy.
 Screening in the first trimester involves the measurement
of PAPP-A (pregnancy associated plasma protein A) &
free b HCG (beta human chorionic gonadotropin) levels in
maternal serum.
 These measurements used in conjunction with USG scanning
that includes assessment of USG markers such as nuchal
translucency (NT) thickness & absence/presence of the nasal
bone (NB) gives a detection rate of abt. 85% while alone the
detection rate with PAPP-A & bHCG was around 65 % 32

33. MATERNAL SERUM SCREENING
 Levels of MSAFP ( alpha Feto protein ), human chorionic
gonadotrophin (HCG) & unconjugated oestriol (UE3) are
measured between 15 & 18 weeks gestation.
 These substances are of fetal origin & cross from the amniotic
fluid into maternal circulation via the placenta.
 Low maternal serum AFP, low UE3 and/or elevated HCG levels
are associated with increased risks of fetal Down syndrome,
 whereas low levels of all three substances suggests increased
risks for trisomy 18 or triploidy.
 High levels of AFP are associated with increased risk of neural
tube & abdominal wall defects;
 while high levels of HCG can be associated with increased risk
for pregnancy complications. 33

34. MATERNAL SERUM SCREENING
 Down So :
1st Trimester Screening Tests
• Maternal Serum Markers
-Preg. asso. Placental Protein A (PAPP-A)
-Free ß hCG
• Fetal Marker- Nuchal thickness
2nd Trimester Screening Tests
• Maternal Serum Markers
-AFP
-E3 Triple test 70%
-hCG Quadruple
-Inhibin A test 34

35. MATERNAL SERUM SCREENING
 Trisomy 18 or Triploidy :
The level of all three substances
( MSAFP, UE3 and HCG ) is low in
trisomy 18
35

36. MATERNAL SERUM SCREENING
 Neural Tube Defects & Abdominal Wall Defects :
• AFP is produced by the yolk sac & later by the liver; it enters the
amniotic fluid & then the maternal serum via fetal urine.
Therefore MSAFP level can be used to determine the AFP levels
from the fetus.
• In the condition of an open NTD (eg, anencephaly, spina bifida)
& abdominal wall defects in the fetus, AFP diffuses rapidly from
exposed fetal tissues into amniotic fluid, and the MSAFP level
rises.
• Also, a NTD can be distinguished from other fetal defects, such
as abdominal wall defects, by the use of an
Acetylcholinesterase test carried out on amniotic fluid. If the
level of acetylcholinesterase rises along with AFP, it is suspected
as a condition of a NTD. 36

37. MATERNAL SERUM SCREENING
• AFP levels are also elevated when the fetus has congenital
nephrosis, or intestinal atresias.
• However, the MSAFP levels also increase with gestational
age, gestational diabetes, twins, pregnancies complicated by
bleeding, & in association with intrauterine growth retardation.
37

38. Separation of fetal cells from the mother's
blood
 A technique currently being developed for clinical use
involves isolating fetal cells from maternal blood to analyse
fetal chromosomes and/or DNA. Ordinarily, only a very
small number of fetal cells enter the maternal circulation;
but once they enter,can be readily identified, they will be
accessible for analysis by a variety of techniques, without
the risks of complications or miscarriage associated with
invasive procedures (CVS & amniocentesis).
 These cells can be collected safely from approximately 12-
18 weeks' gestation onward.
 Nucleated fetal red blood cells (erythroblasts) are currently
the ideal candidates for analysis, although leucocytes &
trophoblast cells may also be identified
38

39. Separation of fetal cells from the mother's
blood
 Fetal blood cells can then be analyzed for the diagnosis of
genetic disorders using FISH, PCR etc.
 Fetal cells separated from a mother's blood have been
successfully used in the diagnosis of cystic fibrosis, sickle
cell anemia, and thalassemia in a fetus.
A. Maternal RBCs B. Fetal RBCs (nucleated) 39

40. INVASIVE TECHNIQUES
Fetal visualization
Fetal tissue sampling
Cytogenetics
Molecular genetics
40

41. INVASIVE TECHNIQUES
Fetal visualization
-Embryoscopy
-Fetoscopy
41

42. Fetal visualization
-Embryoscopy
 Embryoscopy is performed in the first trimester.
 In this technique, a rigid endoscope is inserted via the
cervix in the space between the amnion and the chorion,
under sterile conditions and ultrasound guidance, to
visualize the embryo for the diagnosis of structural
malformations.
42

43. Fetal visualization
-Embryoscopy
43

44. Fetal visualization
-Fetoscopy
 Fetoscopy is performed during the second trimester (after
16 weeks’ gestation).
 In this technique, a fine-caliber endoscope is inserted into
the amniotic cavity through a small maternal abdominal
incision, under sterile conditions and ultrasound
guidance, for the visualization of the embryo to detect the
presence of subtle structural abnormalities.
 It also is used for fetal blood and tissue sampling.
 Fetoscopy is associated with a 3-5% risk of miscarriage;
44

45. Fetal visualization
-Fetoscopy
45

46. Fetal Tissue Sampling
 Amniocentesis
 Chorionic villus sampling (CVS)
 Percutaneous umbilical blood sampling (PUBS)
 Percutaneous skin biopsy
 Other organ biopsies, including muscle & liver
biopsy
46

47. Fetal Tissue Sampling
 Amniocentesis :
 Amniocentesis is an invasive, well-established, safe,
reliable, & accurate procedure & can be performed at 10-
14 weeks of gestation (early amniocentesis) but usually
done at 16-18 weeks of gestation.
 Although early amniocentesis is ass. with a pregnancy loss
rate of 1 – 2 % & an increased incidence of clubfoot.
 It is performed under ultrasound guidance.
 A 22-gauge needle is passed through the mother's lower
abdomen into the amniotic cavity inside the uterus, & 10-
20 mL of amniotic fluid ( that is replaced by fetus within
24hrs ) that contains cells from amnion, fetal skin, fetal
lungs, and urinary tract epithelium are collected. 47

48. Fetal Tissue Sampling
 Amniocentesis :
48

49. Fetal Tissue Sampling
 Amniocentesis :
1. The Cells are grown in culture for chromosomal,
biochemical, & molecular biologic analyses.
2. The Supernatant amniotic fluid is used for the
measurement of substances such as AFP,
AChE,bilirubin & pulmonary surfactant
3. In the third trimester of pregnancy, the amniotic fluid
can be analyzed for determination of fetal lung
maturity.
 The results of cytogenetic and biochemical studies on
amniotic cell cultures are more than 90% accurate.
 Risks with amniocentesis are rare but include 0.5-1.0%
fetal loss and maternal Rh sensitization. 49

50. Fetal Tissue Sampling
 Chorionic villus sampling (CVS) :
 Under USG guidance, a sample of placental tissue is obtained
through a catheter places either transcervically or
transabdominally.
 Performed at or after 10 wks’ gestation,CVS provides the
earliest possible detection of a genetically abnormal fetus
through analysis of trophoblast cells.
 Transabdominal CVS can also be used as late as the 3rd trimester
when amniotic fluid is not available or when fetal blood
sampling cannot be performed.
 CVS, if preformed before 10 wks’ gestation , can be ass. with an
increased risk of fetal limb reduction defects & oromandibular
malformations.
50

51. Fetal Tissue Sampling
 Chorionic villus sampling (CVS) : Transabdominal
51

52. Fetal Tissue Sampling
 Chorionic villus sampling (CVS) : Transcervical
52

53. Fetal Tissue Sampling
 Chorionic villus sampling (CVS) :
 Direct preparations of rapidly dividing cytotrophoblasts can be
prepared, making a full karyotype analysis available in 2 days.
Although direct preparation minimize maternal cell
contamination, most centers also analyse cultured trophoblast
cells, which are embryologically closer to the fetus.This
procedure takes 8 – 12 days.
 In approximately 2% of CVS samples, both karyotypically normal
& abnormal cells are identified.B’coz CVS acquired cells reflect
placental constitution, in these cases, amniocentesis is typically
performed as a followup study to analyze fetal
cells.Approximatally 1/3rd of CVS mosaicisms are confirmed in
the fetus through amniocentesis.
53

54. Fetal Tissue Sampling
 Percutaneous umbilical blood sampling (PUBS)
(cordocentesis)
 PUBS is preformed under USG guidance from the
2nd trimester until term.
 PUBS can provide diagnostic samples for
cytogenetic, hematologic, immunologic, or DNA studies: it can
also provide access for treatment in utero.
 An anterior placenta facilitates obtaining a sample close to the
cord insertion site at the placenta.
 Fetal sedation is usually not needed.
 PUBS has a 1% - 2% risk of fetal loss, along with complication
that can lead to a preterm delivery in another 5%.
54

55. Fetal Tissue Sampling
 Percutaneous umbilical blood sampling (PUBS)
(cordocentesis)
55

56. Fetal Tissue Sampling
 Percutaneous skin biopsy (Preimplantation Biopsy
or Preimplantation Genetic Diagnosis) :
 The most frequent candidates are parents with family histories
of serious monogenic disorders & translocations, who are
therefore at increased risk for transmitting these conditions to
future generations.
 Polar body & blastomere testing are the two primary methods
of PGD.
 In Polar Body Testing, positive test results in two polar bodies
ensure that the egg itself is unaffected – therefore, the
mutation has segregated to the polar body, not to the
developing ovum. Once an egg is found to be unaffected, it is
fertilized via traditional in vitro fertilization (IVF) & implanted
into the uterus. 56

57. Fetal Tissue Sampling
 Percutaneous skin biopsy (Preimplantation Biopsy
or Preimplantation Genetic Diagnosis) :
 Blastomere PGD first requires traditional in vitro fertilization,
after which cells are grown to the 8-cell stage. One or two cells
are harvested & analysed, & an unaffected blastocyst is
implanted into the uterus.
 An advantage to preconception testing over traditional
postconception prenatal diagnosis is that it allows parents to
avoid the possibility of receiving abnormal prenatal diagnosis
results, & thus the difficult decisions associated with pregnancy
management and/or maintenance.
 PGD can be laborious, time-consuming & expensive.
Complicating factors include a high rate of polyspermia, & a
small amount of DNA in polar bodies (making it difficult to
amplify) which can produce less definitive test results. 57

58. Fetal Tissue Sampling
 Other organ biopsies, including muscle & liver
biopsy :
 Fetal liver biopsy is best performed between 17-20 weeks'
gestation under ultrasound guidance.
 Fetal liver biopsy is needed to diagnose inborn errors of
metabolism, such as glucose-6-phosphatase deficiency ,
glycogen storage disease type IA & nonketotic
hyperglycemia.
 Fetal muscle biopsy is carried out under ultrasound
guidance at about 18 weeks' gestation to analyze the
muscle fibers histochemically for prenatal diagnosis of
Becker-Duchenne muscular dystrophy.
58

59. Cytogenetic Investigations
 Chromosome Analysis ( Karyotype Analysis )
 Fluorescence in situ Hybridization (FISH)
59

60. Cytogenetic Investigations
 Chromosome Analysis :
 Chromosome analysis is a technique used to identify
aneuploidy, microdeletions, microduplications & major
structural aberrations.
 The most common method of detecting aneuploidy is
karyotype analysis, wherein metaphase cells are examined
microscopically & the number of chromosomes counted.
 Typically 10–15 cells are analysed to rule aneuploidy in or
out.
60

61. Cytogenetic Investigations
 Chromosome Analysis
 Karyotype Analysis : Each chromosome pair has a unique
banding pattern that can be seen with various stains.
 The most common method of karyotype analysis is Giemsa (G)
banding, wherein chromosomes are denatured (with
trypsin), revealing a pattern of light & dark bands.
 Counting the number of staining chromosomes allows for
detection of aneuploidies.
 Analysing for the absence, presence, rearrangement, etc. of
these bands allows for detection of larger
deletions, duplications and structural aberrations.
 Although G banding is typically used first to analyse prenatal
specimens, various other banding techniques (including
quinacrine (Q), reverse (R), centromeric heterochromatin (C) &
high-resolution banding) may be used to analyse different
portions of particular chromosomes.
61

62. Cytogenetic Investigations
• Chromosome Analysis ( Karyotype Analysis )
Normal Karyotype
62

63. Cytogenetic Investigations
• Chromosome Analysis ( Karyotype Analysis )
Down Syndrome Karyotpe
63

64. Cytogenetic Investigations
 Fluorescence in situ Hybridization (FISH) :
64

65. Cytogenetic Investigations
 Fluorescence in situ Hybridization (FISH) :
 FISH is mainly used to detect the presence or absence of
microdeletions, microduplications & aneuploidy without the full
effort associated with DNA sequencing or complete karyotype
analysis
 This three-step technique allows specific DNA sequences or
chromosomes to be visualized microscopically.
1. A specific, single-stranded DNA probe is hybridized to its
complementary, target DNA sequence, while the cell is in
prophase, metaphase or interphase;
2. fluorescent antibodies are then hybridized to the probe DNA
sequence;
3. finally, the fluorescent signals are examined under the
microscope.
65

66. Cytogenetic Investigations
 Fluorescence in situ Hybridization (FISH) :
66

67. Cytogenetic Investigations
 Fluorescence in situ Hybridization (FISH) :
Human chromosome 1 painting probe (green)
Terminal band painting probe centromere probe
67

68. Cytogenetic Investigations
 Fluorescence in situ Hybridization (FISH) :
 FISH analysis for common aneuploidies (involving chromosomes 13,
18, 21, X and Y) is often performed by simultaneously applying
specific multicoloured centromeric probes. In fetal trisomies, three
probes are present for a specific chromosome, while monosomies
show only one.
(a).A nucleus has been hybridized with probes for chromosomes 18 (aqua), X (green) and one
Y(red).(b) A nucleus has been hybridized with probes for chromosomes 13 (green) and 21 68
(red).

69. Cytogenetic Investigations
 Fluorescence in situ Hybridization (FISH) :
Microdeletions/microduplications detectable by fluorescence in situ
hybridization (FISH)
DISORDER CHROMOSOMAL BAND FINDING
Angelman syndrome 15q12(maternal) Microdeletion
Duchenne Muscular Dystrphy Xp21 Microdeletion
Prader-Willi Syndrome 15q12(paternal) Microdeletion
Retinoblastoma 13q14 Microdeletion
α- thalassemia 16p13 Microdeltion
WAGR syndrome 11q13 Microdeletion
69

70. Molecular genetics
 Direct DNA Analysis
 Linkage Analysis(indirect DNA analysis)
 DNA Sequencing
70

71. Molecular genetics
 Direct DNA analysis :
• Direct mutation analysis involves analysing a target segment
of DNA for the presence of a specific mutation. Like FISH, it
requires knowledge of the correct sequence for the specific
gene or DNA segment before analysis. Once known, the
sample sequence may be compared to the
known, ‘model’, genomic sequence in a variety of
methods, as described below :
 Mutation analysis with restriction enzymes.
 Sequencing of restriction enzyme products.
 Allele-Specific Oligonucleotide (ASO) analysis.
71

72. Molecular genetics
 Direct DNA analysis :
 Mutation analysis with restriction enzymes :
If the putative mutation is known to alter the recognition for
a splice site, direct analysis by restriction enzyme assay is
possible. The presence of a mutation can be detected by
digesting control and sample DNA with the same restriction
enzymes (known to cut the DNA at a specific splice site) and
then analysing resultant DNA fragments (called Restriction
Fragment Length Polymorphisms, or RFLPs ) for differences
by Southern blotting. Those segments containing mutation(s)
at or near a splice site are identifiable because they were not
cut by a restriction enzyme, and are therefore
longer, appearing higher on the Southern blot gel. (Longer
fragments do not migrate as quickly or as far as shorter
fragments.) This technique is used in genetic testing for sickle
cell anaemia. 72

73. Mutation analysis with restriction enzymes :
73

74. Molecular genetics
 Direct DNA analysis :
 Sequencing of restriction enzyme products :
• Disorders secondary to deletion of DNA ( e. g α thalassemia, DMD, CF
& growth harmone deficiency) can be detected by the altered size of
DNA segments produced following a Polymerase Chain Reaction(PCR)
• DNA sequences that have been cut with restriction enzymes can also
be sequenced by a specialized amplification technique. Copies of a
particular piece of DNA(cut by restriction enzymes) are placed into
four vials & amplified by polymerase chain reaction (PCR).
• Fragments from the vials are then allowed to migrate, in parallel,
down a Southern blot gel.
• The shortest fragments travel furthest, the longer segments remain
closer to the top. From top to bottom, the banding pattern produced
represents fragments that decrease in size by one nucleotide base.
The DNA sequence can therefore be read from the shortest, single-
base strand at the bottom of the gel, up to the entire sequence
length at the top 74

75. Molecular genetics
 Direct DNA analysis :
 Allele-Specific Oligonucleotide (ASO) analysis :
• Direct detection of a DNA mutation can also be accomplished by
allele specific oligonucleotide analysis.
• If the PCR –amplified DNA is not altered in size by deletion or
insertion, recognition of mutated DNA sequence can occur by
hybridization with the known mutant allele.
• ASO analysis allows direct DNA diagnosis of Tay-Sachs Disease,
alpha & beta thalassemia, Cystic fibrosis & phenylketonuria
75

76. Molecular genetics
 Linkage Analysis(indirect DNA analysis) :
• Linkage analysis is a means of indirectly detecting a patient’s
mutation status, when several family members are known to be
affected with the same genetic disorder, & when an exact
mutation is not known.
• DNA from affected & unaffected family members is analysed for
polymorphisms such as microsatellite repeats, restriction
fragment length polymorphisms (RFLPs) and variable number
tandem repeats (VNTRs)
76

77. Molecular genetics
 DNA Sequencing :
• DNA sequencing for many disorders has revealed that a
multitude of different mutations within a gene can result in
same clinical disease.
• For Example,Cystic Fibrosis can result from more than 1,000
different mutations.
• Therefore, for any specific disease, prenatal diagnosis by DNA
testing may require both Direct & Indirect methods.
77

78. ®PRENATAL TREATMENT
• In the most situations the diagnosis of prenatal
abnormalities has a subsequent option of
termination of the pregnancy.
• While this applies in most situations, there is
cautious optimism that with the advent of gene
therapy prenatal diagnosis will, in time, lead to
effective treatment in utero.
• Example-Treatment of the autosomal recessive
disorder-Congenital Adrenal Hyperplasia (CAH).
Affected female are born with virilisation of the
external genitalia. There is an evidence that this can
be prevented by powerful steroid therapy at early
gestational age. 78

79. ETHICS & LEGAL ASPECTS
• Before screening or testing pregnancies for underlying genetic
disorders, it is important to consider the ethics of a given
situation. Genetic diagnosis may affect decisions about
maintaining or ending a pregnancy, counselling should be done
in such cases.
• Prenatal diagnosis in high risk cases should be done at
appropriate gestational age as according to MTP act
pregnancies can be terminated only upto 20 weeks of gestation
when Substantial risk of physical or mental abnormalities in the
fetus as to render it seriously handicapped.
79

80. SUMMARY
80

81. 81

82. THANK YOU 82