Cytogenetics Prelim Notes

Walther Flemming
 An Austrian cytologist and professor of anatomy, who published the first
illustrations of human chromosomes in 1882.
 Also referred to the stainable portion of the nucleus as chromatin and first used the
term mitosis
Waldeyer
 Introduced the word chromosome, from the Greek words for “colored body”
Sutton and Boveri
 Formally developed a “chromosome theory of inheritance”
Sutton
 Combined the disciplines of cytology and genetics when he referred to the study of
chromosomes as cytogenetics.
Von Winiwarter
 Concluded that men have 47 chromosomes and women have 48
Painter
 Studied (meiotic) chromosomes derived from the testicles of several men who had
been incarcerated, castrated, and ultimately hanged in the Texas State Insane
Asylum.
 Definitively reported the human diploid chromosome number to be 48 (double the
24 bivalents he saw), even though, 2 years earlier, he had preliminarily reported
that some of his better samples produced a diploid number of 46
 Proposed the X and Y sex chromo- some mechanism in man.
Levitsky
 Formulated the term karyotype to refer to the ordered arrangement of
chromosomes
Hsu
 Reported that, rather than depending on histologic sections, examination of
chromosomes could be facilitated if one studied cells grown with tissue culture
techniques published by Fisher
 Demonstrated the value of this method by using it to examine human embryonic cell
cultures, from which he produced both mitotic metaphase drawings and an
ideogram of all 48 human chromosomes
Hsu and Pomerat
 Realizing the poten- tial of this fortuitous event, reported a “hypotonic shock”
procedure
Ford and Hamerton
 Had modified this technique and had also worked out a method for pretreating cells
grown in culture with colchicine so as to destroy the mitotic spindle apparatus and
thus accumulate dividing cells in the metaphase
Joe Hin Tjio
 An American-born Indonesian, learned about these proce- dures and worked with
Hamerton and Ford to further improve upon them.
Tjio and Levan
 Cytogenetics Prelim Notes

 Optimized the colchicine/hypotonic method for these cells, and in January 1956
(after care- fully reviewing images from decades of previously reported work), they
diplomatically reported that the human diploid chromosome number appeared to
be 46, not 48
Waardenburg
 Made the suggestion that Down syndrome could perhaps be the result of a
chromosomal aberration
Lejeune
 Studied the chromosomes of fibro- blast cultures from patients with Down
syndrome
 Had proved Waardenburg’s hypothesis by reporting the first example of a
chromosomal syndrome in man
Lejeune and colleagues
 Described an extra chromosome in each of these cells
Ford et al.
 Reported that females with Turner syndrome have 45 chromosomes, apparently
with a single X chromosome and no Y
Jacobs and Strong
 Demonstrated that men with Klinefelter syndrome have 47 chromosomes, with the
additional chromosome belonging to the group that contained the X chromosome
Jacobs
 A female with sexual dysfunction was also shown to have 47 chromosomes and was
believed to have an XXX sex chromosome complement
Murray Barr
 Was studying fatigue in repeatedly stimulated neural cells of the cat
Lyon
 Developed the single active X chromosome mechanism of X-dosage compensation in
mammals
 Lyon Hypothesis
Patau et al.
 Described two similar infants with an extra “D group” chromosome who had
multiple anomalies quite different from those seen in Down syndrome
 Trisomy 13
Edwards et al.
 Described “A New Trisomic Syndrome” in an infant girl with yet another
constellation of phenotypic abnormalities and a different autosomal trisomy
 Trisomy 18
Nowell and Hungerford
 Reported the presence of the “Philadelphia chromosome” in chronic myelogenous
leukemia, demonstrating, for the first time, an association between chromo- somes
and cancer
 Cytogenetics Prelim Notes

Lejeune et al.
 Reported that three infants with the cri du chat (“cat cry”) syndrome of phenotypic
anomalies, which includes severe mental retardation and a characteristic kitten-like
mewing cry, had a deletion of the short arm of a B-group chromosome, designated
as chromosome 5.
Jacobs et al.
 Described “aggressive behavior, mental subnormality and the XYY male”, and the
chromosomal instabilities associated with Bloom syndrome and Fanconi anemia
were reported
Nowell
 Observed that the kidney bean extract phytohemagglutinin, used to separate red
and white blood cells, stimulated lymphocytes to divide
 He introduced its use as a mitogen, permitting a peripheral blood sample to be used
for chromosome analysis.
Torbjorn Caspersson
 Observed that when plant chromosomes were stained with fluorescent quinacrine
compounds, they did not fluoresce uniformly, but rather produced a series of bright
and dull areas across the length of each chromosome.
Caspersson and colleagues
 Turned their attention from plants to the study of human chromosomes.
 Hypothesized that the quinacrine derivative quinacrine mustard (QM) would
preferen- tially bind to guanine residues
Caspersson and co-workers
 Had successfully produced and reported a unique “banding” pattern for each human
chromosome pair
Drets and Shaw
 Described a method of pro- ducing similar chromosomal banding patterns using an
alkali and saline pretreatment followed by staining with Giemsa

Cytogenetics Terminologies

 Karyotype – refer to the ordered arrangement of chromosomes

 Trisomy 21 – Down Syndrome
 Trisomy 13 – Patau’s Syndrome; D-trisomy
 Trisomy 21 – Edward’s Syndrome; E-trisomy
 Turner’s Syndrome – have 45 chromosomes, apparently with a single X
chromosome and no Y.
 Klinefelter’s Syndrome – have 47 chromosomes, with the additional chromosome
belonging to the group that contained the X chromosome.
 Female with Sexual Dysfunction – have 47 chromosomes and was believed to
have an XXX sex chromosome complement.
 Barr Body – X chromatin
 Philadelphia chromosome – chronic myelogenous leukemia, an association
between chromosomes and cancer.
 Cytogenetics Prelim Notes

 Cri du chat (“cat cry”) syndrome – phenotypic anomalies, which includes severe
mental retardation and a characteristic kitten-like mewing cry, had a deletion of the
short arm of a B-group chromosome, designated as chromosome 5.
 Phytohemagglutinin – used to separate red and white blood cells, stimulated
lymphocytes to divide.
 Giemsa – a compound developed for identification, in blood smears, of the
protozoan that causes malaria.
 FISH – Fluorescence in situ hybridization

CHAPTER 2: DNA, Chromosomes, and Cell Division

NUCLEIC ACIDS. Write DNA if the phrase describes DNA and RNA if a description of RNA is
given
1. Used as template in replication
2. Used as template in reverse transcription
3. Primer is a short fragment of this
4. The product of a replisome
5. Made up of covalently linked dNTPs
6. Consist of A=T and G=C base pairs
7. Consist of G=C and A=U base pairs
8. Regarded as a gene product
9. Occurs frequently as double stranded
10. Occurs frequently as single stranded

CENTRAL DOGMA OF CELL BIOLOGY. The following word or groups of words are all
associated with the central dogma of cell biology.
Write RP if closely associated with Replication
TC if closely associated with Transcription
TL if closely associated with Translation

1. Primer attaches to the DNA

2. Aminoacyl tRNA
3. DNA exonuclease activity
4. Promoter attaches to the RNA
5. Sealing of nicks in the lagging strand
6. Splicing of introns
7. Semiconservative process
8. 5’ – 7’ methyl guanosine capping
9. Poly-A-tail forms sticky ends
10. Ribosomal assembly
11. Topoisomerase untangles the DNA
12. Joining of Okazaki fragments
13. Interactions among the rRNA, mRNA, and tRNA
 Cytogenetics Prelim Notes

14. TATA box

15. Rho factor terminates the process
16. Addition of UTP to the growing polymer
17. Replisome machinery
18. Attachment of amino acid to the 3’ end of the acceptor arm
19. Occurs in the cytosol
20. Gene products pass through the nuclear pores

Terms on Replication

 Gene – sequence of dNTPs coding for a specific trait

 DNA – sequence of dNTPs serving as template in replication
 RNA – sequence of rNTPs serving as transcript for proteins
 Protein – sequence of amino acids; product of translation
 Gene products – products derived from DNA
 Replication – process of making two double-stranded DNA from a double-stranded
DNA template
 Transcription – process of making single-stranded RNA from a DNA strand
 Reverse Transcription – process of producing DNA from and RNA template
 Translation – process of producing protein as mediated by mRNA
 Plasmid – the circular DNA material in bacteria
 Semi-conservative – described to a new DNA that has an old DNA strand
 Replication bubble – site in DNA consisting of two replication forks
 Helicase – enzyme that cleaves H-bonds between strands of DNA
 dNTPs – the building blocks of a DNA molecule
 DNA polymerases – enzymes which adds dNTPs during replication
 Topoisomerase – enzyme responsible for coiling and uncoiling of DNA
 Leading strand – the single-stranded DNA with continuous addition of dNTPs
 Lagging strand – the single-stranded DNA with discontinuous dNTP addition
 Primase – enzyme responsible for attaching a primer to single-stranded DNA
 Primer – RNA oligonucleotide signaling the start of replication
 Okazaki fragment – oligonucleotides added to the lagging DNA strand
 DNA ligase – enzyme which repairs nicks in the lagging strand
 Replisome – the DNA replicating machine
 oriC – the origin of DNA replication in prokaryotes
 Single-stranded binding proteins – protects single-stranded DNA from attack by
nucleases

Terminology Part II
 Transcription – the synthesis of RNA from a DNA template
 RNA polymerase I – the enzyme that synthesizes the ribosomal precursor RNA
 RNA polymerase II – the enzyme that synthesizes mRNAs
 Promoter – the DNA fragment where the RNA polymerase binds
 Cytogenetics Prelim Notes

 Hogness box – also known as the TATA box in eukaryotic transcription

 tRNA – covalently attaches an amino acid to the ribosome
 L-shaped – the tertiary structure of the tRNA
 mRNA – carrier of the genetic code from DNA to the ribosome
 rRNA – major component and 2/3 of the gross weight of the ribosome
 Closed promoter – the complex formed by the double-stranded DNA, promoter and
RNA polymerase
 Rho factor – the protein that terminates the transcription process
 Exons – the coding regions of the mRNA
 Introns – the non-coding regions of the mRNA
 hnRNA – the RNA composed of introns and exons
 Mature mRNA – the RNA, purely exons, with 7-methyl guanosine cap and poly A tail
 Acceptor arm – the arm in tRNA where the amino acid binds
 Triplet codon – a sequence of three bases specifying an amino acid
 Stop codons – the collective term for the triplets UAA, UAG, and UGA
 Methionine – the first amino acid formed in every translation
 Shine-Dalgarno – the GGAGGU sequence preceding the initiation site for translation
in prokaryote

Terminology Part III

1. The molecule that bears the genetic information found in chromosomes
2. The sugar found in the RNA and DNA backbone
3. The building blocks of nucleic acids
4. Described as a ribonucleotide that triggers the replication process
5. Watson, Crick, Franklin, and _______________ contributed to DNA structure elucidation
6. Noncovalent _____________ Bonds are found between bases in a double-stranded DNA
7. The covalent chemical bonds that link alternating sugar and phosphate in DNA
8. Chargaff discovered that DNA has equal amounts of adenine and ___________
9. There is ________________ base pairing the maintains the double helical DNA structure
10. The replication bubble is made up of two replication ____________
11. The enzyme that helps helicase by unwinding the DNA strands is the _____________
12. The RNA polymerase II synthesizes the ________________ ribonucleic acid
13. Small segments of nucleotides called ______________fragments are formed in the
lagging strand
14. Exons are the ____________ regions of an immature transcript of mRNA
15. The product of semiconservative process of replication is one _______________ DNA
strand and a new synthesized strand of DNA
16. The RNA _____________ is the one that attaches to the promoter during transcription
17. Splicing of _____________ is necessary for the production of a mature mRNA
18. Leder, Khorana and _____________ determined the 3 nitrogen bases constituting a
codon.
19. _____________ Codons out of the 64 represent specific amino acids
20. There are _____________ codons that serve as termination signals of transcription
 Cytogenetics Prelim Notes

21. The ______________ of the DNA code is best explained by the ability of one mRNA codon
to represent two or more amino acids.
22. The nucleotide triplets found on the tRNA molecules are called ______________
23. The _______________ read mRNA transcript one codon at a time
24. The _______________ bond connects the amino acids in a protein molecule
25. The _______________ is made of a single continuous molecule of DNA complexed with
histone and nonhistone proteins
26. The octameric protein core in a nucleosome is made of pairs of histone proteins
H2A, H3, H4, and ____________
27. The 30-nm coil of 6 nucleosomes constitute the ____________
28. The site of protein synthesis is the ____________
29. Chromosomes are at their most condensed state during the mitotic _____________
30. The chromosomes are made up of sister chromatids, telomere, centromere, and
________________________.

Terminologies Part IV
(Parts of Chromosomes)

Centromere  Joins sister chromatids together

 Chromosomal part that interacts with mitotic spindle during
cell division
 Basis for classifying chromosomes into metacentric,
arcocentric, and submetacentric
 Where the kinetochore apparatus is greatly associated with
Telomere  The far end portions of chromosomes
 Protective caps of chromosomal ends
 Plays a role in synapsis during cell meiosis
 Shortening of this structure is associated with tumor-
suppressor mechanism
 Contains repeats of TTAGGG sequence
 Its length decreases with aging and repeated cell division and
cell culture
Nucleolar  Contained in the satellite stalks of arcocentric chromosomes
Organizer Regions  Sites of ribosomal RNA synthesis
 Contain rRNA genes and 10 in number in humans
a-satelite DNA  Tandem array of million base-pair repeats of 171-bp sequence
 Its 17 base-pair portion binds CENP-B, a centromere protein
 Satelite DNA that is located in heterochromatin and associated
with centromeres
Minisatelite  Few thousand base-pairs with 20-70 bp repeats
 Useful markers for gene mapping and identity testing
 Satellite DNA with close similarity with microsatellites
SINES  Short dispersed repetitive DNA
 Cytogenetics Prelim Notes

 Includes the Alu sequence high in G-C content

 Dispersed repetitive DNA made of 90-500 bp associated with
nuclear pre-mRNA and introns of mRNA
LINES  Long dispersed repetitive DNA
 Includes the L1 sequence with high A-T content
 Dispersed repetitive DNA that can be made of 7000 bases

(Types of Chromatin)

Euchromatin  Described as loosely organized, extended and coiled

 Microscopically seen as lightly staining nuclear bodies with GTG
 Regarded as the early replicating chromatin
 Contains active replicating genes

Heterochromatin  Classed into facultative and constitutive

 Described as inactive, highly contracted and late replicating
 Found in the distal end of the Y chromosome
 Associated with the inactive X chromosome
 Microscopically seen as darkly staining nuclear bodies with GTG

Both Euchromatin  Found in eukaryotic cells

and
Heterochromatin

(Types of Cell Division)

Mitosis  Two daughter cells are produced per cycle

 Involves all somatic cells
 Occurs throughout the entire lifetime
 Parent and daughter cells are both haploid
 For the purpose of growth and tissue repair
 Exhibited by organism during asexual reproduction
 Characterized by four stages
 The parent cell undergoes only one division

Meiosis  Made up entirely of two divisions

 Daughter cells produced are four in number
 For sexual reproduction
 Daughter cells are genetically different
 Production of haploid daughter cells
 Involves the gametes (sperm and ovum)
 Completed only during sexual maturity
 Cytogenetics Prelim Notes

(Cell Cycle)

Gap 1  The 1st stage of interphase

 Chromosomes exist as single chromatids
 Cells are metabolically active during this stage
 Longest stage of the cell cycle

Gap 2  Interphase stage that lasts for 3 hours

 Endstage of interphase
 Interphase stage preparatory for mitosis

Mitosis  Consists of four stages resulting in daughter cells

 Lasts for 1-2 hours

Synthesis  Production of two identical sister chromatids

 Occurs only for 5 hours
 Stage where replication happens
 The stage that directly follows the G1

Cytokinesis  Stage that follows the mitotic stage

Gap 0  Cells are not dividing during this stage; arrested stage

(Stage of Meiosis)

Leptotene  Chromosomes begin to condensed (2n)

 First stage of prophase I

Zygotene  Synapsis happens

Pachytene  Synapsis completed

 Where crossing-over happens between bivalents

Diplotene  In males, the X and Y chromosomes associate end to end

 Chromosomes thicken and shorten further and homologous
chromosomes begin to repel
 Chromosomes are held together via charismata

Diakinesis  Chromosomes reach their greatest contraction during this last

stage of prophase.

Metaphase I  Disappearance of the nuclear membrane and formation of the

meiotic spindle.
 The bivalents line up on the equatorial plate with their
 Cytogenetics Prelim Notes

centromeres randomly oriented toward opposite poles.

Anaphase I  Centromeres of each bivalent separate and migrate to opposite

poles

Telophase I  The two haploid sets of chromosomes reach opposite poles and
the cytoplasm divides.
 The result is two cells containing 23 chromosomes, each
composed of two chromatids.

Metaphase II  1st stage of meiosis II

 Directly follows telophase I
 23 chromosomes line up on the equatorial plate

Anaphase II  Chromatids separate and move to opposite poles

Telophase II  Net result is four cells with 23 chromatids

 Stage where cytokinesis occurs during meiosis II

Chapter 3: Human Chromosome Nomenclature

Terminologies Part V

 FISH – fluorescence in situ hybridization

 GTG – G-bands by Trypsin and Giemsa
 ISCN – International System of Human Cytogenetic Nomenclature
 CGH – Comparative Genomic Hybridization
 “p” in the p arm – Petite
 Dr. Mary Lyon – proponent of Lyon hypothesis
 Lyon hypothesis – states that during early development , one of the x chromosomes
in a female gets turned off and this is maintained in all descendant cells of the clone.
 “Dosage compensation” – it is a mechanism to equalize the dosage of X-
chromosome gene product by means of inactivating one of female X-chromosome in
mammals.
 Sperm – gamete that bears either X or Y chromosomes
 Phenotypic sex – refers to an individual’s sex as determined by their
internal/external genitalia, characteristics and behavior.

Gonadal Sex of an Infant with a sex compliment of:

 XX – Female
 XY – Male
 XXX – Female
 XXY – Male
 XYY – Male
 Cytogenetics Prelim Notes

Chromosomal Group (Old Nomenclature)

Chromosome # Description (Based on the
centromere location)
Group A 1-3 1 & 3 – large metacentrics
2 – large submetacentrics
Group B 4-5 Large submetacentrics (almost
indistinguishable)
Group C 6-12 + X Middle size metacentrics
Group D 13-15 Middle size acrocentrics with NOR
Group E 16-18 16 – short metacentric
17 & 18 – short submetacentrics
Group F 19-20 Short metacentrics
Group G 21-22 + Y Short acrocentrics with NOR,
except Y