Gene Therapy, Gametogenesis, Prenatal genetic testing

Question 1

Aptamere:

Answer 1

Aptamere: RNA molecules that specifically bind to a small molecule or a protein – they are selected from a random pool.

Question 2

How can we treat a disease with gene therapy?

Answer 2

Nucleic acid selection

Question 3

Nucleic acid selection - DNA Methods Examples

Answer 3

Plasmid
Linear DNA (viral)
Artificial chromosome

Question 4

Nucleic acid selection - RNA Methods Examples

Answer 4

Antisense RNA
Ribozyme
Silencing RNA
microRNA (miRNA)
Aptamere

Question 5

How to introduce the nucleic acid?

Answer 5

Selection of vector - Most commnly viral

Question 6

Episomal:

Answer 6

Episomal: A segment of DNA that can exist and replicate either autonomously in the cytoplasm or as part of a chromosome, mainly found in bacteria.

Question 7

What are examples of Viruses used in Gene therapy as vectors?

Answer 7

Adenovirus, AAV, Lentivirus

Question 8

Spermatogonium and Oogonium:

What is the Chromosomal Content? (2n/n)

Answer 8

Spermatogonium and Oogonium are:

Diploids (2n)

Question 9

Primary Spermatocyte and Oocyte :

What is the Chromosomal Content? (2n/n)

Answer 9

Primary Spermatocyte and Oocyte are:

Diploids (2n)

Question 10

Secondary Spermatocyte and Oocyte :

What is the Chromosomal Content? (2n/n)

Answer 10

Secondary Spermatocyte and Oocyte are:

Haploids (n)

Question 11

Spermatid and Ootid :

What is the Chromosomal Content? (2n/n)

Answer 11

Spermatid and Ootid are :

Haploids (n)

Question 12

Spermatozoa and Ovum:

What is the Chromosomal Content? (2n/n)

Answer 12

Spermatozoa and Ovum are :

Haploids (n)

Question 13

What is the name of the process that allows Spermatogonium and Oogonium to become Primary Spermatocyte and Oocyte?

Answer 13

Maturation

NOT MEIOSIS

Question 14

What is the name of the process that allows Primary Spermatocyte and Oocyte to become Secondary Spermatocyte and Oocyte?

Answer 14

MEIOSIS I

Question 15

What is the name of the process that allows Secondary Spermatocyte and Oocyte to become Spermatid and Ootid?

Answer 15

MEIOSIS II (Completed only in Fertilization)

Question 16

What is the definition of Infertility:

Answer 16

Infertility: inability to conceive after 12 months. ~10% couple are infertile.

Question 17

Rare genetic causes of non obstructive azoospermia (low sperm count) – all account for <5% of infertility: (4)

Answer 17

− Kallmann syndrome
− Robertsonian translocations
− XX males (pseudohermaphroditism)
− Point mutations / CNV

Question 18

When is In Vitro Fertilization and Embryo Transfer (IVF + ET) indicated for patients?

Answer 18

Infertile female (+ healthy sperm)

Question 19

When is Intracellular Sperm Injection (ICSI) indicated for patients?

Answer 19

Infertile male with abnormal sperm

Question 20

Preimplantation Genetic Diagnosis (PGD):

Answer 20

Preimplantation Genetic Diagnosis (PGD):
Determine the genotype to avoid
transmission of genetic disease
(monogenic disease)

Question 21

Preimplantation Genetic Screening (PGS):

Answer 21

Preimplantation Genetic Screening (PGS):
Determine the karyotype to avoid
transmission of genetic disease
(numerical chromosomal aberrations)

Question 22

What are three main tissues for Tissues for Preimplantation Biopsy?

Answer 22

➢ Polar body – from egg stage
➢ Blastomere – from cleavage stage
➢ Trophectoderm – from blastocyst stage

Question 23

What is used for HLA Typing as a sample?

Answer 23

➢ Blastomere – from cleavage stage

➢ Trophectoderm – from blastocyst

Question 24

What is used for PGS and PGD as a sample?

Answer 24

➢ Polar body – from egg stage
➢ Blastomere – from cleavage stage
➢ Trophectoderm – from blastocyst

Question 25

What is the disadvantage of using Polar body for Preimplantation Biopsy?

Answer 25

Maternal only

Question 26

What is the disadvantage of using Blastomere for Preimplantation Biopsy?

Answer 26

Mosaicism is common

Question 27

What is the disadvantage of using Trophectoderm for Preimplantation Biopsy?

Answer 27

Not all embryos reach blastocyst stage the same day

Question 28

Clinical Indications for Using PGD/PGS:

Answer 28

* Single gene defects (family history of inherited disorder) * Balanced translocations in one of the parents * Advanced maternal age (aneuploidy) * Repetitive IVF failure * Recurrent pregnancy loss * Embryo selection (HLA matching siblings)

Question 29

Molecular Genetic Techniques used in PGS:

Answer 29

* Karyotyping with G-band * FISH * MLPA * aCGH * NGS (New Genome Sequencing) * QF-PCR (Quantitative Fluorescence)

Question 30

What is the gold standard for PGS Method of testing?

Answer 30

• Karyotyping with G-band

Question 31

array-Comparative Genomic Hybridization (aCGH): What is it?

Answer 31

A technique used to detect CNVs. Whole genome of two different fluorescent color labelled and hybridized to slide fixed healthy metaphase chromosomes. fluorescent fragments are bound to chromosomes depending on the number of copies rate of fluorescent signal should be equal (= 1)

Question 32

array-Comparative Genomic Hybridization (aCGH): What Does it mean if the signal is more than 1.2?

Answer 32

tested samples the genome fragments give a more intense signal - tested section of the genome has EXTRA copies

Question 33

array-Comparative Genomic Hybridization (aCGH): What Does it mean if the signal is less than 0.85?

Answer 33

tested samples of genome fragments signal decreased - the tested section of the genome indicates a DEFICIT.

Question 34

Non-invasive Fetal Trisomy (NIFTY) test:

Answer 34

from week 10th, Allow to check for trisomy and other chromosomal aberration of chromosome: 9, 13, 16, 18, 21, 22, X, Y

Question 35

Clinical use of aCGH:

Answer 35

• Disease focused arrays (DMD array) – Haematological and cancer arrays • Prenatal arrays • Preimplantation arrays

Question 36

aCGH - Advantages compared to karyotyping:

Answer 36

– Easily noticeable differences | – Higher resolution (approx. 100 kb) than the resolution of banding techniques (5-10 Mb)

Question 37

aCGH - Advantages compared to FISH:

Answer 37

Not only specific, pre-selected regions can be examined

Question 38

aCGH - Disadvantage:

Answer 38

Contamination (in tumor samples are healthy cells as well).