RFLP= Restriction fragment length polymorphism

Slides:



Advertisements
Presentasi serupa
2. Introduction to Algorithm and Programming
Advertisements

Common Effect Model.
Validitas & Reliabilitas
Game Theory Purdianta, ST., MT..
K-Map Using different rules and properties in Boolean algebra can simplify Boolean equations May involve many of rules / properties during simplification.
OLEH SUDRAJAT FMIPA UNMUL Struktur Sel Bakteri.
BAGIAN III Lapisan Data Link.
Presented By : Group 2. A solution of an equation in two variables of the form. Ax + By = C and Ax + By + C = 0 A and B are not both zero, is an ordered.
Pertemuan 05 Sebaran Peubah Acak Diskrit
GENE THERAPY AN INTRODUCTION Agustina Setiawati, M.Sc., Apt.
Menulis Kolom  Kolom adalah opini atau artikel. Tidak seperti editorial, kolom memiliki byline.  Kolom Biasanya ditulis reguler. Biasanya mingguan atau.
1 Pertemuan 09 Kebutuhan Sistem Matakuliah: T0234 / Sistem Informasi Geografis Tahun: 2005 Versi: 01/revisi 1.
Ruang Contoh dan Peluang Pertemuan 05
Teknologi dna rekombinan
Masalah Transportasi II (Transportation Problem II)
1 Pertemuan 21 Function Matakuliah: M0086/Analisis dan Perancangan Sistem Informasi Tahun: 2005 Versi: 5.
BAB 6 KOMBINATORIAL DAN PELUANG DISKRIT. KOMBINATORIAL (COMBINATORIC) : ADALAH CABANG MATEMATIKA YANG MEMPELAJARI PENGATURAN OBJEK- OBJEK. ADALAH CABANG.
PERTEMUAN KE-6 UNIFIED MODELLING LANGUAGE (UML) (Part 2)
Pertemuan 07 Peluang Beberapa Sebaran Khusus Peubah Acak Kontinu
HAMPIRAN NUMERIK SOLUSI PERSAMAAN NIRLANJAR Pertemuan 3
Verb Tense Tense denotes the time of the action indicated by a verb. The time is not always the same as that indicated by the name of the tense.
Population Genetics: Selection and mutation as mechanisms of evolution
1 Pertemuan 11 Function dari System Matakuliah: M0446/Analisa dan Perancangan Sistem Informasi Tahun: 2005 Versi: 0/0.
9.3 Geometric Sequences and Series. Objective To find specified terms and the common ratio in a geometric sequence. To find the partial sum of a geometric.
By: Vandrektus Derek,A.Md
How to fix/make sinking office chair reusable Post by:
DISTRIBUSI BINOMIAL.
STATISTIKA CHATPER 4 (Perhitungan Dispersi (Sebaran))
HTML BASIC (Contd…..) PERTEMUAN KEDUA.
Pert. 16. Menyimak lingkungan IS/IT saat ini
MENDEL LAW AND ITS DEVIATIONS
Cartesian coordinates in two dimensions
Cartesian coordinates in two dimensions
REKAYASA GENETIKA.
Pengujian Hipotesis (I) Pertemuan 11
Matakuliah : I0014 / Biostatistika Tahun : 2005 Versi : V1 / R1
DISTRIBUSI BINOMIAL.
Metabolisme asam nukleat II
Pemrograman Berorientasi Objek
Pendugaan Parameter (II) Pertemuan 10
REAL NUMBERS EKSPONENT NUMBERS.
Open and Closed Social Stratification
Manajemen Proyek Perangkat Lunak (MPPL)
Master data Management
Perhatikan gambar berikut ini baik-baik,
Analisis Korelasi dan Regresi Berganda Manajemen Informasi Kesehatan
A SMALL TRUTH TO MAKE LIFE 100%. Hard Work H+A+R+D+W+O+R+K = 98% Knowledge K+N+O+W+L+E+D+G+E = 96%
How to Set Up AT&T on MS Outlook ATT is a multinational company headquartered in Texas. ATT services are used by many people widely across.
Is it different ? HEREDITY SUBSTANCES HEREDITY SUBSTANCES.
How You Can Make Your Fleet Insurance London Claims Letter.
Things You Need to Know Before Running on the Beach.
How to Pitch an Event
Don’t Forget to Avail the Timely Offers with Uber
Suhandi Wiratama. Before I begin this presentation, I want to thank Mr. Abe first. He taught me many things about CorelDRAW. He also guided me when I.
Simultaneous Linear Equations
Take a look at these photos.... Also, in case you're wondering where this hotel is, it isn't a hotel at all. It is a house! It's owned by the family of.
THE INFORMATION ABOUT HEALTH INSURANCE IN AUSTRALIA.
Group 3 About causal Conjunction Member : 1. Ahmad Fandia R. S.(01) 2. Hesti Rahayu(13) 3. Intan Nuraini(16) 4. Putri Nur J. (27) Class: XI Science 5.
Lesson 2-1 Conditional Statements 1 Lesson 2-1 Conditional Statements.
MAKING A CONCLUSION AND RECOMMENDATION
By Yulius Suprianto Macroeconomics | 02 Maret 2019 Chapter-5: The Standard of Living Over Time and A Cross Countries Source: http//
Metabolisme asam nukleat II
Right, indonesia is a wonderful country who rich in power energy not only in term of number but also diversity. Energy needs in indonesia are increasingly.
Website: Website Technologies.
Vector. A VECTOR can describe anything that has both MAGNITUDE and DIRECTION The MAGNITUDE describes the size of the vector. The DIRECTION tells you where.
"More Than Words" Saying I love you, Is not the words, I want to hear from you, It's not that I want you, Not to say but if you only knew, How easy, it.
A SHORT ESSAY OF CIVIL ENGINEERING BY : ALFATIHATU RAHMI CIVIL ENGINEERING ENGINEERING FACULTY ANDALAS UNIVERSITY PADANG.
Draw a picture that shows where the knife, fork, spoon, and napkin are placed in a table setting.
Wednesday/ September,  There are lots of problems with trade ◦ There may be some ways that some governments can make things better by intervening.
Transcript presentasi:

RFLP= Restriction fragment length polymorphism RFLP analysis RFLP= Restriction fragment length polymorphism Refers to variation in restriction sites between individuals in a population Acu pada variasi di (dalam) lokasi pembatasan antar[a] individu di (dalam) suatu populasi These are extremely useful and valuable for geneticists (and lawyers) Ini [yang] berharga dan bermanfaat untuk ahli genetika ( dan para pengacara) On average two individuals (humans) vary at 1 in 1000 bp Pada [atas] rata-rata dua individu ( manusia) bertukar-tukar pada 1 1000 bp The human genome is 3x109 bp Manusia [itu] Genome adalah 3x109 bp This means that they will differ in more than 3 million bp. [Alat/ makna] ini yang mereka akan berbeda lebih dari 3 juta bp By chance these changes will create or destroy the recognition sites for Restriction enzymes Kebetulan perubahan ini akan menciptakan atau menghancurkan lokasi pengenalan [itu] untuk Enzim Pembatasan

RFLP Lets generate a restriction map for a region of human X-chromosome Biarkan menghasilkan suatu peta pembatasan untuk suatu daerah manusia X-Chromosome The restriction map in the same region of the X chromosome of a second individual may appear as Pembatasan memetakan di [dalam] daerah sama X chromosome suatu detik/second individu boleh nampak [ketika;seperti] Normal GAATTC Mutant GAGTTC

RFLP The internal EcoRI site is missing in the second individual Ecori Yang internal Lokasi adalah hilang individu yang kedua For X1 the sequence at this site is GAATTC CTTAAG This is the sequence recognized by EcoRI The equivalent site in the X2 individual is mutated GAGTTC CTCAAG Now if we examine a large number of humans at this site we may find that 25% possess the EcoRI site and 75% lack this site. Sekarang jika kita menguji sejumlah besar manusia pada lokasi ini [yang] kita boleh temukan bahwa 25% menguasai Ecori [itu] Lokasi dan 75% kekurangan lokasi ini We can say that a restriction fragment length polymorphism exits in this region Kita dapat kata[kan bahwa suatu panjangnya fragmen pembatasan polymorphism pergi daerah ini These polymorphisms usually do not have any phenotypic consequences Silent mutations that do not alter the protein sequence because of redundancy in Codon usage, localization to introns or non-genic regions or do not affect protein Structure/function. Ini polymorphisms pada umumnya tidak mempunyai manapun konsekwensi phenotypic Mutasi diam yang tidak mengubah urutan protein [itu] oleh karena pemborosan di (dalam) Codon Pemakaian, Lokalisasi ke introns atau daerah non-genic atau tidak mempengaruhi protein Structure/Function

RFLP RFLP are identified by southern blots In the region of the human X chromosome, two forms of the X-chromosome are Segregating in the population. Digest DNA with EcoRI and probe with probe1 What do we get?

RFLP Digesting with BamHI and performing Southern blots with the above probe produces the following results for X1/X1, X1/X2 and X2/X2 individuals: Pencernaan dengan Bamhi dan melakukan/menyelenggarakan Selatan menodai dengan di atas pemeriksaan menghasilkan hasil yang berikut untuk X1/X1, X1/X2 Dan X2/X2 Individu: There is no variation with respect to the BamHI sites, all individuals produce the same banding patterns on Southern blots Tidak ada variasi berkenaan dengan Bamhi Lokasi, semua individu menghasilkan yang sama menjilid pola teladan pada [atas] Titik noda Selatan If we used probe2 for southern blots with a BamHI digest what would be the Results for X1/X1, X1/X2 and X2/X2 individuals? Jika kita probe2 digunakan untuk selatan menodai dengan suatu Bamhi mencerna apa [yang] akan adalah Hasil untuk X1/X1, X1/X2 Dan X2/X2 Individu? If we used probe2 for southern blots with a EcoRI digest what would be the results for X1/X1, X1/X2 and X2/X2 individuals? Jika kita probe2 digunakan untuk selatan menodai dengan suatu Ecori mencerna apa [yang] akan adalah hasil untuk X1/X1, X1/X2 Dan X2/X2 Individu?

RFLP RFLP’s are found by trial and error and they require an appropriate probe and enzyme They are very valuable because they can be used just like any other genetic marker to map genes RFLP’S ditemukan oleh mencoba-coba dan mereka memerlukan suatu enzim dan pemeriksaan sesuai Mereka adalah sangat bernilai sebab mereka dapat digunakan seperti halnya penanda [yang] hal azas keturunan lain untuk memetakan gen They are employed in recombination analysis (mapping) in the same way as conventional Allelic variants are employed Mereka dipekerjakan analisa penggabungan-ulang ( pemetaan) dengan cara yang sama seperti Varian [yang] Allelic konvensional dipekerjakan The presence of a specific restriction site at a specific locus on one chromosome and its absence at a specific locus on another chromosome can be viewed as two allelic forms of a gene Kehadiran suatu lokasi pembatasan spesifik pada suatu tempat spesifik pada [atas] satu chromosome dan ketidakhadiran nya pada suatu tempat spesifik pada [atas] chromosome lain dapat dipandang sebagai dua format [yang] allelic suatu gen The phenotype in this case is a Southern blot rather than white eye/red eye Phenotype dalam hal ini adalah suatu Selatan menodai dibanding/bukannya mata eye/red putih Lets review standard mapping: To map any two genes with respect to one another, they must be heterozygous at both loci. Biarkan standard tinjauan ulang [yang] memetakan: Untuk memetakan manapun dua gen berkenaan dengan satu sama lain, mereka harus heterozygous pada kedua-duanya loci.

Mapping Gene W and B are responsible for wing and bristle development Centromere Telomere To find the map distance between these two genes we need allelic variants at each locus W=wings B=Bristles w= No wings b= no bristles To measure genetic distance between these two genes, the double heterozygote is crossed to the double homozygote

B b Mapping Both the normal and mutant alleles of gene B (B and b) are sequenced and we find W B Centromere Telomere B 2 3 E b 5 GAATTC AAATTC By chance, this mutation disrupts the amino acid sequence and also a EcoRI site! If DNA is isolated from B/B, B/b and b/b individuals, cut with EcoRI and probed in A Southern blot, the pattern that we will obtain will be B/B Bristle B/b Bristle b/b No bristle

Mapping Therefore in the previous cross (WB/wb x wb/wb), the genotype at the B locus can be distinguished either by the presence and absence of bristles or Southern blots WB/wb x wb/wb Female Male Wings No wings Bristles No Bristles Southern blot: Southern blot: 5 and 2 kb band 5 kb band There are some phenotypes for specific genes that are very painful to measure Having a RFLP makes the problem easier

Mapping The same southern blot method can be employed for the (W) wing Locus with a different restriction enzyme (BamHI) if an RFLP exists at this locus !! You make the DNA, digest half with EcoRI and probe with bristle probe Digest the other half with BamHI and probe with the wing probe. W 8 GTATCC B B w 4 4 GGATCC B B B

Mapping To find the map distance between genes, multiple alleles are required. We can determine the distance between W and B by the classical Method because multiple alleles exist at each locus (W & w, B & b) You find a new gene C. There are no variants of this gene that alter the phenotype of the fly, that you can observe. Say we don’t even know the function of this gene. You can’t even predict its phenotype. However the researcher identified an RFLP variant in this gene. Centromere Telomere W B C R

Mapping C c 8 E 2 6 With this RFLP, the C gene can be mapped with respect to other genes: Genotype/phenotype relationships for the W and C genes WW and Ww = Red eyes ww = white eyes CC = 8kb band C/c = 8, 6, 2 kb bands cc = 6, 2 kb bands To determine map distance between R and C, the following cross is performed W C w c ------------ ------------ w c w c

Mapping W B C R w W C(8) c(6,2) w c(6,2) w c(6,2) Male gamete (wc) Female gamete

SNPs, RFLPs, point mutations GAATTC GAGTTC GACTTC Pt mut SNP RFLP

PCR If a region of DNA has already been cloned and sequenced, the sequence can be used to isolate and amplify that sequence from other individuals in a population. Jika suatu daerah DNA telah sequenced dan cloned, urutan dapat digunakan untuk mengisolasikan dan memperkuat suara urutan itu dari individu lainnya di (dalam) suatu populasi. Individuals with mutations in p53 are at risk for colon cancer Individu dengan mutasi di (dalam) p53 berhadapan dengan resiko untuk kanker tanda titik dua To determine if an individual had such a mutation, prior to PCR One would have to clone the gene from the individual of interest (construct a genomic library, screen the library, isolate the Clone and sequence the gene). Untuk menentukan jika perorangan telah mutasi seperti itu, sebelum PCR Orang akan harus clone gen dari individu [bunga/minat] ( membangun suatu perpustakaan genomic, menyaring perpustakaan [itu], mengisolasikan Clone [itu] Dan Urutan gen). With PCR, the gene can be isolated directly from DNA isolated from that individual. No lengthy cloning procedure Only small amounts of genomic DNA required 30 rounds of amplification can give you >109 copies of a gene

Genotype and Haplotype In the most basic sense, a haplotype is a “haploid genotype”. Haplotype: particular pattern of sequential SNPs (or alleles) found on a single chromosome in a single individual The DNA sequence of any two people is 99 percent identical. Sets of nearby SNPs on the same chromosome are inherited in blocks. Blocks may contain a large number of SNPs, but a few SNPs are enough to uniquely identify the haplotypes in a block. The HapMap is a map of these blocks and the specific SNPs that identify the haplotypes are called tag SNPs. Haplotyping: involves grouping individuals by haplotypes, or particular patterns of sequential SNPs, on a single chromosome. Microarrays, and sequencing are used to accomplish haplotyping.

SNP mapping is used to narrow down the known physical location of mutations to a single gene. The human genome sequence provided us with the list of many of the parts that make a human. The HapMap provides us with indicators which we can focus on in looking for genes involved in common disease. Using the HapMap data we compare the SNP patterns of people affected by a disease with those of unaffected people. This allows researchers to survey the whole genome quickly and identify genetic contributions to common diseases--the HapMap Project has simplified the search for gene variants.

A recessive disease pedigree

Mapping recessive disease genes with DNA markers DNA markers are mapped evenly across the genome The markers are polymorphic- they look slightly different in Different individuals. We can tell looking at a particular individual which grandparent Contributed a certain part of its DNA. If we knew that grandparent carried the disease, we could say That part of the DNA might be responsible for the disease. A B C D E F G H I 4 different alleles at each locus A1, A2, A3, A4 B1, B2, B3, B4 C1, C2,………….

Mapping recessive disease genes with DNA markers B C D E F G H I Grandparents 1 and 4 and offspring 1 and 4 have a disease We would look at the markers and see that ONLY at position G do offspring 1 and 4 have the DNA from grandparents 1 and 4. It is therefore likely that the disease gene will be somewhere near marker G.

Genetic polymorphism Genetic Polymorphism: A difference in DNA sequence among individuals, groups, or populations. Genetic Mutation: A change in the nucleotide sequence of a DNA molecule. Genetic mutations are a kind of genetic polymorphism. Genetic Variation Single nucleotide Polymorphism (point mutation) Repeat heterogeneity

Repeats Variation between people- small DNA change – a single nucleotide polymorphism [SNP] – in a target site, RFLPs and SNPs are proof of variation at the DNA level, Satellite sequences: a short sequence of DNA repeated many times in a row. Chr1 Interspersed Chr2 tandem

Repeats Satellite sequences: a short sequence of DNA repeated many times in a row. 2 5 6 E E E Chr1 Interspersed Chr2 E E E tandem 3 1 4 0.5 5 3 1 Repeat probe

Repeat expansion Tandem repeats expand and contract during recombination. Mistakes in pairing leads to changes in tandem repeat numbers E 4 1 2 Individual 1 E E 2 Individual 2 E E 3