The Ugly

Lets Think About This!

How big is the human genome?

About 3 billion bases per haplotype Or 6 billion bases in a diploid cell

Estimated ~30,000 genes
Comprised of 2% of our genome Rest made up of non-coding sequences or so called junk DNA but..

How many genes do we have?

1 kilobase (Kb) is 1,000 bases 1 megabase (Mb) is 1,000,000 bases or one thousand kilobases or 1000 x 1000.

1 million bases of sequencing data is equivalent to megabyte of computer data (RAW DATA!!)
Does not include other annotations or other information.

3 billion nucleotide is equivalent to gigabytes of computer data and full diploid sequence is 1.5 gigabytes.

So How Big Are Our Genes?

Big genes are over 500 Kbs pre-mRNA

200 or so genes have been identified so far

Mainly found in CNS related developmental sites Chromosomal instabilities sites

Average gene size 25-30 kb Genes can be as small as a single exon, e.g. histone genes or as big as the dystrophin gene with 79 exons.

Exon size 200 bp average

However
Exon 26 of the apoB gene (APOB) is 7.6 kb Exon 15 of the adenomatous polyposis coli gene (APC) 6.5 kb Exon 11 of the BRCA1 breast cancer gene 3.4 kb

Intron size 0.5 kb to 30 kb mRNA size 2.5 kb 5 UTR 100 bases Coding DNA 1.5-1.8 kb or 500-600 codons 3 UTR 600-800 bases

Note could be underestimated because many long 3 UTR hasnt been reported.

Chromosome abnormalities can be defined as changes resulting in a visible alteration of the chromosomes.

Banding pattern can reveal structures that are 1-10 Mb regions Smallest loss or gain of material visible by traditional methods on standard cytogenetics is ~4 megabases of DNA 100-200 Kb probes Fiber FISH few Kbs Genomic copy number differences 5-10 Kbs

Fluorescent in situ hybridization

Array comparative genomic hybridization (aCGH)

High-resolution CGH arrays can detect structural variations at 200 bp Oligonucleotide Arrays or DNA arrays uses 25 nucleotide length probes

Types of Chromosomal Abnormality

Constitutional abnormality

Sperm or Egg Early embryo post fertilization

Somatic or acquired abnormality Two categories

Structural Numerical

Causes of Aneuploidy
Nondisjunction: failure of paired chromosomes to separate (disjoin) in anaphase of meiosis I, or failure of sister chromatids to disjoin at either meiosis II or at mitosis. Nondisjunction in meiosis produces gametes with 22 or 24 chromosomes, which after fertilization by a normal gamete make a trisomic or monosomic zygote. Nondisjunction in mitosis produces a mosaic. Anaphase lag: failure of a chromosome or chromatid to be incorporated into one of the daughter nuclei following cell division, as a result of delayed movement (lagging) during anaphase. Chromosomes that do not enter a daughter cell nucleus are lost.

Mixoploidy
Mixoploidy includes mosaicism (an individual possesses two or more genetically different cell lines all derived from a single zygote) and chimerism (an individual has two or more genetically different cell lines originating from different zygotes). Abnormalities that would be lethal in constitutional form may be compatible with life in mosaics.

BALANCED TRANSLOCATIONS
Involving 2 chromosomes 46,XX,t(8;14)(q24;q12)
Involving 3 chromosomes 46,XX,t(8;22;14)(q24;q11;q12)

RECIPROCAL TRANSLOCATION

4 2 4 2 2

Balanced translocation

UNBALANCED TRANSLOCATIONS
Robertsonian Translocations 45,XX,der(14;21)(q10;q10) 45,XX,rob(14;21)(q10;q10) for constitutional studies only

Dicentric Chromosomes 45,XY,dic(2;4)(p12;q14)

ROBERTSONIAN TRANSLOCATIONS
Chromosomal rearrangement between the 5 acrocentric chromosome pairs

13 14 15 21 22

ROBERTSONIAN TRANSLOCATIONS

21 21 14 14

ROBERTSONIAN TRANSLOCATIONS

DERIVATIVE CHROMSOME
Generated by more than one aberration within a single chromosome or rearrangement involving two or more chromosomes: 46,XX,der(7)add(7)(p22)del(7)(q22q34)
46,XY,der(9)inv(9)(p11q12)t(9;22)(q34;q11.2), der(22)t(9;22)

NONRECIPROCAL TRANSLOCATION

4 del 2 2 4 2 der 4

45,XY,der(7)t(7;17)(q22;q11.2),-17

DICENTRIC CHROMOSOMES

4 dic(2;4) 2 4 2

acentric fragment (2;4)

DICENTRIC

This karyotype is a Robertsonian Translocation. In this case the entire q, or long arm, plus centromere of a chromosome 13 has been fused with the entire q arm, plus centromere of a chromosome 14. This particular example is unbalanced and results in trisomy 13. There are two normal chromosome 13's plus the chromosome 13 involved with the translocation, thus there are three copies of chromosome 13. The translocation is shown as the right chromosome 14 in the karyotype. The karyotype is written as: 46,XY,+13,dic(13;14)(p11.2;p11.2).

46: the total number of chromosomes. The chromosome number remains 46 because the long arms of chromosomes 13 and 14 have basically fused into one chromosome. XY: the sex chromosomes (male). +13: indicates the presence of an extra chromosome 13.

dic(13;14): dicentric chromosome involving chromosomes 13 and 14. As with many Robertsonian translocations, the centromeres of both chromosomes are present, thus the "dicentric" designation.
(p11.2;p11.2): breakpoints in chromosomes 13 (p11.2), and 14 (p11.2) respectively.

INSERTIONS
Direct 46,XY,ins(5)(q14q21q31) intra 46,XY,ins(5;1)(q14;q22q32) inter

Inverted 46,XY,ins(5)(q14q31q21) 46,XY,ins(5;1)(q14;q32q22)

INSERSION

DELETION

del(2)

DELETIONS

Terminal Deletions 46,XX,del(7)(p11) Interstitial Deletions 46,XX,del(5)(q13q33)

PERICENTRIC INVERSION
A B C D E A C B D E F

PARACENTRIC INVERSION
A B D C E

A B C D E

46,XY,inv(9)(p12q13)

DUPLICATIONS
A B C D E A B C D C D E A

or

B D C C D E

dir dup

inv dup

DUPLICATIONS

Direct Duplications 46,XX,dup(2)(q22q25) Indirect Duplications 46,XX,dup(2)(q25q22)

ISOCHROMOSOMES

RING CHROMOSOME

ISOCHROMOSOMES AND RINGS

Isochromosomes 46,XX,i(17)(q10) Rings 46,XX,r(4)(p14q31) Ring with unknown origin 46,XX,r(4)

RING

FRAGILE SITES
FRAGILE X

Female 46,X,fra(X)(q27.3) Male 46,Y,fra(X)(q27.3)

MOSAICISM
45,X[6]/47,XXX[3]/46,XX[15] 48,XY,+8,+9[6]/46,XY[14] Actual number of cells given in brackets, not ratios or percentage The normal clone is given last In case of multiple abnormal clones, the largest clone is given first Mosaicism vs Chimera

mos 45,X[12]/46,XX[20]

chi 46,XX[20]/46,XY[10]

In chimera, the largest clone is given first

Clone evolution

Clone evolution

Clone evolution

COMPOSITE KARYOTPE
Great karyotypic heterogeneity within tumor population.

Common changes but no two cells share the same structural and numerical aberrations:

Composite Karyotype: (cp)

45,XX,-15,del(17)(q11.1) 46,XX,+7,-15,del(17)(q11.1) 46,XX,+12,-15 47,XX,+7 47,XX,+15,del(17)(q11.1) 48,XX,+12,+15 Composite karyotype will be: 45~48,XX,+7,+12,+15,-15,del(17)(q11.1)[cp6]

Ploidy Range
23+/- 11= 12 - 34 haploid range 46+/- 11= 35 - 57 diploid range 69+/- 11= 58 - 80 triploid range 92+/- 11= 81 - 103 tetraploid range Basic formula: xn +/- 11 x = ploidy level; n= haploid #

FISH Nomenclature
ish: in situ hybridization- metaphase FISH
nuc ish: nuclear ish - interphase FISH pcp: partial chromosome paint wcp: whole chromosome paint

46,XX.ish 9(WCP9x2)

abl/bcr Genes on Diploid Cells and Ph Positive CML Cells