draft sooca case 4 2014.pdf

8/10/2019 Draft Sooca Case 4 2014.pdf

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CASE REVIEW

Tata, 2 YO, Boy

CC: Looked Very Pale

HT PE LE

Looked very pale had

for 6 months

This similar health

happens to both

family members of

father and mother

They receive blood

transfusion reguraly

Immunization (+)

Sick (-)

Nutritional problem (-)

Chronic bleeding (-)

Screened

abnormalities (-)

Looks pale (+)

Conjunctiva anemis

(+)

Sclera yellowish (+)

Abdomen

hepatosplenomegaly

(+)

Haemoglobin

(normal)

His parents are

carriers (from

screening) -

thalassemia

Mutation in -globin

chain (+)

Diagnosis: -Thalassemia

Treatment:

Pharmacology: -

Non-Pharma: Routine blood transfussion


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FINAL CONCEPT

, 2 YO

Mutation In Gene Expression

Parents Carrier

Gene Exp &

Gene Tech

Mutation

Failure In mRNA Splicing

Nucleic Acid,

DNA, RNA, &

Protein Syntesys

Mutation -Globin Chains

Imbalance Of -Globin Chains

-Thalassemia MayorBasic Diagnosys

and Clinicall Signs

Free -Chains

Agregat -ChainsErythroblast Destroyed

(Ineffective

Erytrhopoesis)

Free Precipitates to

Damage RBC

Abnormal Structure

Intravascular

Hemolysys

Erytrocyte Hb

RBC Life Spen

Spleen Destroys RBC

RBC

RBC Production

Erythropoetic Tissues (In

Liver & Spleen) / Work HardPaleAnemia Heme


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Hepatosplenomegaly Biliverdin

Unconjugat

ed Bilirubin

Icteric

Sclera

Treatment BHP & PHOP


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-THALASSEMIA MAYOR

1.

Explain the structure, function, resources and utility of nucleic

acid

2.

Explain the DNA organization, replication and repair

3.Explain the RNA structure, synthesis and processing

4.Explain protein synthesis and genetic code

5.

Explain the regulation of gene expression

6.Explain the gene technology and genetic examination for

diagnosis

7.

Explain the definition of gene, chromosom, and mutation

8.BHP: Ethical issue in frequent blood transfusion

9.PHOP: Premarital screening


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NUCLEIC ACID

1. Structure of Nucleic Acid

Each nucleotide is composed of:

Pentose sugar

o Deoxyribose (DNA)without oxygen atom on carbon-2

o Ribose (RNA)


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Nitrogen Bases

o Purin (Body : Adenine, Guanine ; Free : Xanthine, Hypoxanthine)

o Pyrimidine (Cytocine, Thymine (DNA), Uracil (RNA))

Phosphate group

o Provided by H3PO4, able to support segments of nucleic acid

2. Function / Utility

Form genetic code


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important component of protein synthesis

Energy source for protein synthesis

Gene therapy and gene technology

Metabolic process

3. Source :

- Amino acids

- Sugar

- Food rich in nucleotides (brain, spinach, anchovy, caviar)

- Nucleotide released in intracellular metabolism


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DNA

Structure

Nucleotide : Consist of 3 major component : pentose sugar, phosphate

group, and nitrogen base

DNAs structure is double helix that coiled around common axis. The chains are

paired in an antiparallel manner, that is, the 5end of one strand is paired with

the 3- end of the other strand

Complementary chain structure of DNA show that the amount of purine and

pyrimidine are the same (A + G = C + T). Adenine will be paired with

thymine by 2 hydrogen bond, while Guanine will be paired with Cytosine by 3

hydrogen bond

Backbone of DNA consist of phosphate-sugar group. Pentose sugar of DNA is 2-

deoxyribose

2 nucleotide joined by phosphodiesther bond on 3rd or 5th C atom Nucleosome : consist of DNA segment that warp around histone octamer.

Nucleosome + H1 histone (histone protein tightly bond to chromatin)

called chromatosome.

Histone protein : protein with + charge that contain may arginine and lysine that

could interact withcharge and phosphate group p(ionic bond). Kind of histone

protein

H1 joined in the solenoid

H2A

H2B

H3

H4

There are three major structural forms of DNA: the B form, described by Watson

and Crick in 1953, the A form, and the Z form. The B form is a right-handed helix

with ten residues per 360 turn of the helix, and with the planes of the bases

perpendicular to the helical axis. The A form is produced by moderately

dehydrating the B form. It is also a right-handed helix, but there are eleven base

pairs per turn. The Z-DNA is a left handed helix that contains about twelve base

pairs per turn.

replication

Type of DNA replication accepted by universal is semiconservative type. The

process are :

Separation of the two complementary DNA strands: DNA strain must first

separate (or melt), at least in a small region. In prokaryote, it begins at a single,

unique nucleotide sequencea site called the origin of replication. In eukaryote,


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replication begins at multiple sites along the DNA helix. Helicase is the enzyme

responsible for separating the double chain on the replication forks on the

recognizable sequence of DNA.

Single strain DNA protein is responsible to prevent the two separated DNA chain

back together

Because of the separation of DNA chain in one side, the other part of DNA starts

getting tight. Topoisomerase is responsible for unwinding tight parts of DNA.

After the DNA chain is separated into 2 part, the synthesis begins. DNA

Polymerase 3 starts to synthesis new-born DNA. The synthesis begins from the 3

end. Because DNA consist of 2 anti-parallel backbone, that means they move in

different direction. So, there are 2 sites :

Leading strain : continuously synthesize from 5 to 3 direction without

distraction

Lagging strain : Because DNA synthesize need 3-OH end, the synthesize

must be initiated by RNA-primase. From that, DNA polymerase 3 start to synthesize newborn DNA. After finished

synthesizing 1 DNA fragment, the Okazaki fragment, RNA primase are removed

by DNA polymerase 1. Then, they will be joined together by ligase enzyme


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DNA repair

Mismatch repair

Base excision repair


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Nucleotide excision repair

Double strand break


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RNA

A. Definition

RNA stands for ribonucleic acid, which is a polymeric molecule made up of one or more

nucleotides. A strand of RNA can be thought of as a chain with a nucleotide at each

chain link. Each nucleotide is made up of a base(adenine, cytosine, guanine, or uracil), a

ribose sugar, and a phosphate.

RNA is divided into four types:

a. rRNA

b. mRNA

c. tRNA

d.

snRNA

B. Structure

A, G, C, U nitrogenous bases

pentose sugar (ribose sugar)

Pphosphate group


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C. Function

D. RNA Synthesis

RNA synthesis is the process of transcribing DNA nucleotide sequence information into

RNA sequence information. It is catalyzed by a large enzyme called RNA polymerase.

In eukaryotic cells it occurs in the cell nucleus, while in prokaryotic cells, it occurs in the

cytoplasm.

There are three steps to this process, which are initiation, elongation, and termination.

Illustration to these steps is shown in the picture on the next page.


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E. RNA Processing


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PROTEIN SYNTHESIS AND GENETIC CODE

Protein synthesis is translating genetic code to make amino acids which will bind each other to

form polypeptides.

The genetic code is a dictionary that identifies the correspondence between a sequence of nucleotidebases and a sequence of amino acids. Each individual word in the code is composed of three nucleotide bases.These genetic words are called codons. Codons are presented in the mRNA language of adenine (A), guanine (G),

cytosine (C), and uracil (U). The four nucleotidebases are used to produce the three-base codons.

Characteristics of genetic code:

1.

Specificity (unambigous), a particular codon always codes for the same amino acid. Example: UUU always

codes for phenylalanine

2.

Universality, the genetic codes virtually universal, same encoding will occur in every synthesis that

happens. An exception occurs in mitochondria, wich a few codons have different meanings.

3.

Degeneracy, a given amino acid may have more than one triplet coding for it. For example, arginine

is specified by six different codons. Only Met and Trp have just one coding triplet.

4.

Nonoverlapping and commaless, the code is read from a fixed starting point as a continuous

sequence of bases, taken three at a time. For example, AGCUGGAUACAU is read as

AGC/UGG/AUA/CAU without any punctuation between the codons.


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GENE EXPRESSION

Influenced by

1. Hormones

2. Heavy metals3. Chemicals

Regulation

a. Positive: Expression of genetic information increased by the presence of specific regulatory

element (activator)

b. Negative: Expression of genetic information diminished by the presence of specific

regulatory element (repressor)

Regulation response

1.

Type A (prokaryotes): An increased extent of gene expression that is dependent upon thecontinued presence of the inducing signal. When the signal is removed, the amount of gene

expression diminishes to its basal level.

2. Type B (during development of organism): Increased amount of gene that is transient even

in the continued presence of the regulatory signal.

2. Type C (during development of differenciated function in a tissue or organ): In response to

the regulatory signal, an increased extent of gene expression that persists indefinitely after

termination of the signal (the signal acts as a trigger)

Regulation of gene expression

Pada prokariot, sebagian besar DNA cetakan dapat dilakukan transkripsi namun untuk

eukariot hrus dilakukan remodeling kromatin (modifikasi kromatin) terlebih dahulu guna

ditranskripsi. Khusus pada prokariot, DNA cetakan itu berupa operon. APA ITU OPERON?

Operon: Gugus gugus cetakan DNA yang diawali dengan promoter, operator dan beberapa

gen yang bersebelahan(operon lac). Guna promoter adalah tempat terdapatnya RNA

polymerase dan operator berfungsi untuk menyalakan dan mematikan(repressor) dalam

proses regulasi.Operator pada prokariot berfungsi sama dengan metilasi DNA pada eukariot.

1. Modifikasi kromatin

Gen dalam kromatin sangat terpadatkan biasanya tidak ditranskripsikan.

Asetilasi histon tampaknya melonggarkan struktur kromatin dan meningkatkan transkripsi.

Metilasi DNA umumnya mengurangi transkripsi.

2. Transkripsi


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Regulasi inisiasi transkripsi: unsur kontrol DNA mengikat faktor transkripsi spesifik.

Penekukan DNA memungkinkan aktivator menyentuh protein di promoter, sehingga

menginisiasi transkripsi.

Regulasi terkoordinasi.

3. Pemrosesan mRNAPenyambungan RNA alternatif

4. Translasi

Inisiasi dari translasi dapat dikontrol melalui regulasi dari faktor inisiasi.

5. Pemrosesan dan degradasi protein.

Oleh proteasom

6. Degradasi mRNA.

Setiap mRNA memiliki panjang usia yang khas, ditentukan sebagian oleh sekuens pada UTR 5

dan 3


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GENE EXAMINATION

RFLP (Restriction Fragment Length Polymorphism)

RFLP analysis is used to identify changes in genetic sequences which occur at a site where the

restriction enzyme cuts.

RFLP is based on the chance of comparing each DNA profile resulted from the cutting of

targeted DNA with the restriction enzyme.

Steps of RFLP:

1.

DNA Isolation

The process of isolating the DNA fragments with extraction and lytic method. Usually the

process is done with the help of homogenization and usage of additional extraction buffer

to prevent DNA from damaging.

Separation of DNA from other cell components is done using the centrifuge.

2.

Cutting of DNA using the restriction enzyme and electrophoresis gel

The result of DNA isolation will then cut using certain restriction enzyme which selected

carefully. Every restriction enzyme which place in a suitable environment will recognize and

slice the DNA to form the fragments. The fragments will then undergo electrophoresis using

the Aragosa gel.

Electrophoresis

The standard lab procedure for separating DNA by size (e.g. length in base pairs) for visualization

and purification.

The principle of the basic categories of analyses used to characterize DNA and RNA:

Electrophoretic Separation

Movement of DNA or RNA in response to an electric field will be proportional to the molecularweight or length of the molecule. This property is used to characterize the size of nucleic acid

fragments by electrophoretic separation.

Hybridization

The process of combining two complementary single-stranded DNA or RNA molecules and

allowing them to form a double-stranded molecule through base pairing.

Hybridization is defined as the interaction between two single-stranded nucleic acid molecules

to form a duplex (double-stranded) molecule.

Principle: based on the complementary base pairing of their respective sequences.

High-stringency conditionshigh temperature (close to Tm ), low salt, and high formamidewill only allow the most perfectly matched duplex structures to remain in a stable helix

conformation.


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GENE TECHNOLOGY

DNA Cloning

DNA cloning is a process of producing the identical DNA from its parental DNA.

Vectors are the agent that carries DNA fragment into the living cell in order to increase the

amount of DNA fragments.

There are a few vectors used within the process:

1.

Bacteria

a.

Plasmid : small circular DNA molecule which have an ability to replicate itself

b.

Phage : viruses that infected the bacteria

c.

Cosmides : a larger fragments of DNA which resulted from the combination of

plasmid and phage

2.

Viral

a.

Adenovirus

b.

Retrovirus3.

Artificial Chromosome

DNA Recombinant

DNA recombinant is the formation of new genetic

combination by inserting a DNA molecule into certain

vector, making it possible for them to integrate and

undergo a replication process within other organism.

Purpose of DNA recombinant:

1.

Gene mapping: determine the specific location of

gene in particular chromosome.2.

Production of protein for research and diagnosis

purposes.

3.

Examination of molecular analysis in disease.

Gene Therapy

Gene therapy is the use ofDNAas adrugto treat disease by delivering therapeutic DNA into a patient's

cells. Mostly involves using DNA that encodes a functional, therapeutic gene to replace amutatedgene.

There are two types of gene therapy, yet only one of which has been used in humans:1.

Somatic Gene Therapy

2.

Germline Gene Therapy

Principle of gene therapy

Inserting precursor cell to the location of the damaged cell so that the precursor cell will have the same

expression with damaged cells and able to replace the function of the damaged cells.
http://en.wikipedia.org/wiki/DNAhttp://en.wikipedia.org/wiki/DNAhttp://en.wikipedia.org/wiki/DNAhttp://en.wikipedia.org/wiki/Biologic_medical_producthttp://en.wikipedia.org/wiki/Biologic_medical_producthttp://en.wikipedia.org/wiki/Biologic_medical_producthttp://en.wikipedia.org/wiki/Mutatehttp://en.wikipedia.org/wiki/Mutatehttp://en.wikipedia.org/wiki/Mutatehttp://en.wikipedia.org/wiki/Mutatehttp://en.wikipedia.org/wiki/Biologic_medical_producthttp://en.wikipedia.org/wiki/DNA


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PCR (Polymerase Chain Reaction)

Technic used in certain DNA fragments amplification in an in vitro mediumusing a pair of

oligonucleotide.

Seperating the double-strand DNA, breaking thehydrogen bond in order to form the single-strand DNA. Temperature needed: 92-94 oC.

Denaturation

Primer forward and reverse is searching for itspair in the DNA strands.Annealing

Taq polymerase replicate the DNA fragment.

Temperature needed: 720CElongation


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MUTATIONDefinition

A mutation is defined as a permanent change in the DNA. Mutatations that affect germ

cells are transmitted to the progeny and can give rise to inherited disease. Mutations

that arise in somatic cells understandably do not cause hereditary diseases but areimportant in the genesis of cancers and some congenital malformations. Human genetic

disorders can be broadly classified into three categories :

1. Disorders related to mutations in single genes with large effects: Highly penetrant,

they usually follow the classic Mendelian pattern of inheritance and are also

referred to as Mendelian disorders.

2. Chromosomal disorder: Uncommon, but with high penetrance

3. Complex multigenic disorders: Often expressed in polymorphisms, although it is often

of small effect and low penetrance unless in huge amounts of polymorphisms

expressed (in multigenic or polygenic conditions as it is termed) Penetrance = the

frequency of manifestation of a hereditary condition in individuals. Mutations mayresult in partial or complete deletion of a gene or affect a single base.

Types of mutations

1. Point mutations

It may alter the code in a triplet of bases and lead to the replacement of one

amino acid by another in the gene product. There are several types of this

mutation

. Missense mutations: Replacement of normal amino acid with a very different

one that can cause some physiological or physical changes (often detrimental).

Eg. -thalassemia due to CTC changed in CAC.

. Nonsense mutations: May cause severe translation errors, causing often severe

physiological or physical effects. Eg. Beta-thalassemia major due to codon for

glutamine (CAG) creates stop codon (UAG), this creates premature termination

of betaglobin gene translation and causes severe form of anemia called o-

thalassemia.

. Silent mutation: No change in protein function can be observed due to the

mutation being silent (has no visible physiological or physical effects on the

mutant subject).

Caused due to base substitution, there are 2 types of base substitution :. Transition: Changes purine to purine / pyrimidine to pyrimidine.

. Transversion: Changes purine to pyrimidine / pyrimidine to purine.

2. F rameshift mutations

It alters the reading frame of the DNA strand, it is usually detrimental for an

organismsfitness. They are caused by :


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BHP & PHOP

BHP

Informing patient about the reason why the patient should get the transfusion blood.

Informing patient about the procedure of blood transfussion.

Informing patient about the risk, benefit, purpose, and requirement of blood transfusion to

the donature and the recepient.

PHOP

Preventive : Doing genetic counseling and genetic screening before married

(premartial screening )

Promotive : Educate and persuading people the importance of genetic counseling

before married to prevent the genetical disease happen

Currative : Giving the patient blood transfusion, stem cell transfusion, and bone

marrow transplant.

Rehabilitative : Doing routine check up to prevent any risk happen after blood

transfusion

draft sooca case 4 2014.pdf

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