INTRODUCTION TO REAL TIME PCR IS GIVEN, basic principle of realtime pcr, along with the process of operating this, diagrammatic representation of the process, advantages and disadvantages o f reatimem pcr, applications of the same is also there
In this lecture tried to introduce some basic methods of DNA sequencing like pyrosequencing, sequencing by ligation, sequencing by synthesis and Ion Semiconductor Sequencing
and describe them. Also introduced some new sequencing method (third generation sequencing) like SMRT (Single Molecule Real-Time Sequencing) and GridION.
Techniques of DNA Extraction, Purification and QuantificationBHUMI GAMETI
Introduction
The overall process…
Uses of isolated genomic DNA
Extraction of DNA from plant material
Components of DNA extraction solutions
Cell Lysis or Cell disruption :
Purification of DNA
CTAB Method
Phenol–chloroform extraction
PROTEINASE K
Salting out
Silica adsorption method
Magnetic beads
FTA Paper
Nucleic acid quantification
Agarose Gel Electrophoresis
UV spectroscopy
DNA quantification using NanoDrop
INTRODUCTION TO REAL TIME PCR IS GIVEN, basic principle of realtime pcr, along with the process of operating this, diagrammatic representation of the process, advantages and disadvantages o f reatimem pcr, applications of the same is also there
In this lecture tried to introduce some basic methods of DNA sequencing like pyrosequencing, sequencing by ligation, sequencing by synthesis and Ion Semiconductor Sequencing
and describe them. Also introduced some new sequencing method (third generation sequencing) like SMRT (Single Molecule Real-Time Sequencing) and GridION.
Techniques of DNA Extraction, Purification and QuantificationBHUMI GAMETI
Introduction
The overall process…
Uses of isolated genomic DNA
Extraction of DNA from plant material
Components of DNA extraction solutions
Cell Lysis or Cell disruption :
Purification of DNA
CTAB Method
Phenol–chloroform extraction
PROTEINASE K
Salting out
Silica adsorption method
Magnetic beads
FTA Paper
Nucleic acid quantification
Agarose Gel Electrophoresis
UV spectroscopy
DNA quantification using NanoDrop
Probe labeling is defined as sequence use to search the mixture of nucleic acid for molecule containing complementary sequence.In molecular biology, hybridization is a phenomenon in which single-stranded deoxyribonucleic acid (DNA) or ribonucleic acid (RNA) molecules anneal to complementary DNA or RNA
Basics of DNA isolation, What is chemistry behind it. Presently the laboratory of animal science department ,Göttingen university using this technique for dna isolation in pig blood sample.
In a detail description of the two major blotting techniques. Right from its history to the result interpretation put forth in a concise way. Helps understand these protocols with ease.
WHAT IS BLOTTING?
Blotting is a technique for detecting any macromolecules that we deal with like DNA, RNA or proteins, which are initially present in a complex mixture.
TYPES OF BLOTTING:
Southern Blotting
Northern Blotting
Western Blotting
NORTHERN BLOTTING
A northern blotting is a laboratory method used to detect specific RNA molecules among a mixture of RNA (mRNA).
The technique was developed in 1979 by James Alwine and his colleagues.
Northern blotting can be used to analyze a sample of RNA from a particular tissue or cell type in order to measure the expression of particular genes.
Northern blotting involves the use of electrophoresis to separate RNA samples by size, and detection with a hybridization probe complementary to part of or the entire target sequence.
The term ‘northern blot’ actually refers specifically to the capillary transfer of RNA from the electrophoresis gel to the blotting membrane. However the entire process is commonly referred to as northern blotting.
PROCEDURE
1.RNA isolation:
2.Separation of RNA using gel electrophoresis:
3.BLOTTING:
4.Hybridization with labelled probe:
5.WASHING OFF EXCESS PROBES
Sanger sequencing is a method of DNA sequencing based on the selective incorporation of chain-terminating dideoxynucleotides by DNA polymerase during in vitro DNA replication.
Definition, principle, Chemical used during the process, application, advantages, and disadvantages of both techniques. along with relevant case study for better understand
Probe labeling is defined as sequence use to search the mixture of nucleic acid for molecule containing complementary sequence.In molecular biology, hybridization is a phenomenon in which single-stranded deoxyribonucleic acid (DNA) or ribonucleic acid (RNA) molecules anneal to complementary DNA or RNA
Basics of DNA isolation, What is chemistry behind it. Presently the laboratory of animal science department ,Göttingen university using this technique for dna isolation in pig blood sample.
In a detail description of the two major blotting techniques. Right from its history to the result interpretation put forth in a concise way. Helps understand these protocols with ease.
WHAT IS BLOTTING?
Blotting is a technique for detecting any macromolecules that we deal with like DNA, RNA or proteins, which are initially present in a complex mixture.
TYPES OF BLOTTING:
Southern Blotting
Northern Blotting
Western Blotting
NORTHERN BLOTTING
A northern blotting is a laboratory method used to detect specific RNA molecules among a mixture of RNA (mRNA).
The technique was developed in 1979 by James Alwine and his colleagues.
Northern blotting can be used to analyze a sample of RNA from a particular tissue or cell type in order to measure the expression of particular genes.
Northern blotting involves the use of electrophoresis to separate RNA samples by size, and detection with a hybridization probe complementary to part of or the entire target sequence.
The term ‘northern blot’ actually refers specifically to the capillary transfer of RNA from the electrophoresis gel to the blotting membrane. However the entire process is commonly referred to as northern blotting.
PROCEDURE
1.RNA isolation:
2.Separation of RNA using gel electrophoresis:
3.BLOTTING:
4.Hybridization with labelled probe:
5.WASHING OFF EXCESS PROBES
Sanger sequencing is a method of DNA sequencing based on the selective incorporation of chain-terminating dideoxynucleotides by DNA polymerase during in vitro DNA replication.
Definition, principle, Chemical used during the process, application, advantages, and disadvantages of both techniques. along with relevant case study for better understand
gives a very brief info about western blotting procedures, attractive slides, with creative animation effects, i hope this ppt of mine works good for seminar and for educational purposes.
ADN et génotypage - Présentation de la 5e édition du Cours international « Atelier Paludisme » - Célestin RAZAFINJATO - Médecin - INSPC - Antananarivo, Madagascar - razafinjatocelestin@yahoo.fr
TECHNIQUES DE BIOLOGIE MOLECULAIRE EN PARASITOLOGIE ET MYCOLOGIE
EXTRACTION D'AN
TECHNIQUE PCR
SEQUENCAGE D'AN
GENOTYPAGE
APPORT DE LA BIOLOGIE MOLECULAIRE EN PARASITOLOGIE ET MYCOLOGIE MEDICALE
Entomologie moléculaire et étude de la structuration génétique des anophèles - Présentation de la 2e édition du Cours international « Atelier Paludisme » - RANDRIANASOLO Laurence - INSTITUT PASTEUR de MADAGASCAR BP 1274 Antananarivo, Madagascar - laurandrianas@yahoo.fr
De l'ADN à la protéine & de l'antigène à l'anticorps - Présentation de la 5e édition du Cours international « Atelier Paludisme » - Edith Christiane BOUGOUMA - Chercheur - Centre National de Recherche sur le Paludisme / CNRFP - Ouagadougou, Burkina Faso - eddycnrfp@yahoo.fr
Titulaire d'un doctorat en physiologie de la nutrition et ancien chercheur, je vous invite ici à découvrir le monde merveilleux de notre ADN et du code génétique. Le personnage et les encadrés pointent les aspects importants à retenir pour le BTS diététique
2. Introduction:
Les blots sont des techniques de transfert d'ADN, d'ARN
et de protéines sur un support de sorte qu'ils peuvent
être séparés
C'est une technique indirecte d'hybridation de l'ADN
mise au point en 1975 par le biochimiste Edwrad
Southern.
La technique est connue comme le transfert d'ADN ou
«Southern blot »
Le Southern est une technique longue qui est la base de
l'essor de la biologie moléculaire mais qui est souvent
remplacée par la PCR.
3. Le but de southern blot:
Elle permet de détecter une séquence
d'ADN génomique dite unique
Cette technique permet de localiser
une séquence d’ADN parmi les
fragments ayant migrés, grâce à une
sonde marquée
4. Principe :
La clé de ce procédé est l'hybridation.
Hybridation: Il est le processus de formation
d'une molécule d'ADN double brin entre une
sonde d'ADN simple brin et un ADN cible
simple brin.
Il ya 2 éléments importants de l'hybridation:
◦ Les réactions sont spécifiques à des sondes vont
seulement se lier à des cibles avec une
séquence complémentaire.
◦ La sonde peut trouver une molécule de cible
dans un mélange de millions de molécules
apparentées mais non complémentaires.
5. Protocole :
Extraction de l’ ADN
Restriction de l’ADN
l’électrophorése
Transfert
Hybridation
autoradiographie
7. EXTRACTION D’ADN:
Traitement par l’ARNase
PouréliminerlesARNs
précipitation de l’ADN
=addition d'éthanol ou d'isopropanol dans la phase aqueuse
déprotéinisationde la solution
=extraction / solvants organiques, en général du phénol +/-
chloroforme
Lyse des cellules ou des tissus
=broyage ou pas + extraction/détergents
15. Champ d’application :
Pour identifier l'ADN spécifique dans un
échantillon d'ADN
Pour isoler l'ADN désiré pour la construction
d'ADNr
Identifier les mutations, délétions et des
réarrangements de gènes
Utilisé dans le pronostic du cancer,EX:MYCN
Le diagnostic de VIH-1 et les maladies
infectieuses
Dans RFLP Restriction Fragment Length
Polymorphism:
Dans empreintes génétiques
le diagnostic prénatal et des maladies
génétiques