What are the three basic steps in PCR technique?

Denaturation Primer annealing Extension and DNA Synthesis From which organism DNA polymerase I enzyme is extracted? Thermus aquaticus What are the different methods to study the PCR products? Gel Electrophoresis Cloning Sequencing What are the different methods to study the PCR products? Gel Electrophoresis Cloning Sequencing What is the formula of annealing temperature? Tm=4(G+C)+2(A+T) While designing a primer what should be its GC content? 50-60% Name any DNA Polymerase enzyme except taq polymerase? Pyrococcus furiosus Which type of PCR is used to quantify gene expression, to screen for mutation & finding SNPs? Real Time PCR What are the four parameters which should be taken care of during primer designing? Primer length Base composition Melting temperature Nature of sequence Name a technique which can be used for diagnosis of molecular disease? FISH or Cytogenetic Test Name a tool for primer designing? Primer 3 Which PCR do not use primers? Colony PCR SOEing is used for? Overlapping extentions How many sets of primers are used in nested PCR? Two ASO stands for? Allele Specific Oligomer hybridization Which promoter is linked with the PCR primer in the PTT? T7 Promoter

What effects are generated due to Stop codons generated by point or frameshift mutations? Premature stop of translation and reduced size of the translated proteins Which are the two oligo nucleotide probes used in OLA? Capture and Reporter Oligos What are the three phases that can be seen in the linear view of PCR? Exponential Linear Plateau What is Q-Gene? Software tool GENETICS QUIZ SUBMITTED BY: GURPREET KAUR What are the characteristics of primer No internal structures specificity no hairpin formation Formula for annealing temperature Tanneal= Tmelting temp of primer-4oC Application of multiplex PCR Genome identification Steps of RT-PCR Denaturation annealing elongation synthesis of cDNA Tools used in real time PCR REST Q-gene qBASE Full form of REST Relative expression software tool Full form of DART Data analysis for real time PCR Types of PCR Hot start PCR Nested PCR Colony PCR Real time PCR Multiplex PCR RT-PCR Which enzyme is used in cold finish PCR New hybrid polymerase Hybridisation based method is used for detection of gene mutation Full form of SSCP Single strand conformation polymorphism Silver staining is used for Staining of low molecular weight proteins

Name a dye used to stain proteins in PAGE Coomassie brilliant blue Which PCR is used for the detection of thalessemia? Real Time-PCR What should be the GC content in primer? 50-60% OLA is used for Detection of SNP A disease detected by OLA OLA Which colour are the fluorescent dyes used in OLA Green and blue Full form of CCM Chemical cleavage method Enzyme used to join capture and reporter oligos? DNA ligase A good primer have melting temperature in the range of 52 C to 65 C A good primer has lack of Dimerization capability The temperature at which half the DNA strands are single stranded and half are double stranded is called as melting temperature What is the internal structure those are avoided by an ideal primers? Hair pin Self dimer dimer Average G+C content in an ideal primer is 50 60% PCR was invented in ---------- by -------------1985 Karry mullis What is the PCR equation Xn = X0 ( 1+ E )n In hybridisation probe technique a florescein dye at ------- end and other with a rhodamine dye at --------- end 3 prime and 5 prime Rt PCR are used for Detection of specific RNA s Which type of PCR are used for the screening the DNA from bacterial colonies? Colony PCR Which type of polymerase are used in colony PCR? Special chimeric DNA polymerase Which type of PCR have successive cycles of lowering annealing temperature 0.5 or 1 C TOUCH DOWN PCR DNA binding dye is?

SYBR green Phase of PCR in graph? Exponential Linear plateau The temperature at which primers anneal to the template DNA is called Annealing temperature What is the full form of PPT? PROTEIN TRANSLATION TEST SNP s stands for? Single nucleotide polymorphism PTT is based on a combination of PCR tanscription and translation Which method is used for SNP genotyping? OLA Oligonucleotide ligation assay DNA fragments of upto ------- can be analysed by CCM 2 Kb RACE stands for? Rapid amplification of c DNA ends What is the full form of SSCP Single strand conformation polymorphism What is the full form of DGGE Denaturing gradient gel electrophoresis What is the full form of RFLP? Restriction fragment length polymorphism The duration of denaturation step in first cycle of PCR is Usually 2 minutes at 94C The success of PCR depends on the use of Correctly designed primers PCR technique was developed by Kary mullis in 1985 The details of PCR techniques and its mechanism are described by Erlich 1989 PCR technolgy book The primer length affects PCR efficiency True The two primers used in PCR must not have regions complementary to each other because this would result in primer dimer formation Two primers used in PCR reaction have identical --------for successful and specific amplification Tm The efficiency of amplification declines with an increase in the Length of target sequence The thermostable enzyme used in PCR is Taq polymerase Name the thre steps used in PCR? Denaturation

Annealing Primer extension The concentration of mg++ is maintained between 1 and 4 mM Allele specific oligomer hybridisation method is based on Dot blot method The modification of RT- PCR is called as RACE (Rapid amplification of c DNA ends) In which type of PCR, a critical component like mg ++ or taq polymerase is left out from the reaction mixture? Hotstart PCR A Temperature higher than the ideal annealing temperature reduces PCR temperature true OLA stands for Oligonucleotides ligation assay Typically ,there are ------- capture probes for each allele two The size of translation product is analysed by Gel electrophoresis Hydroxylamine reacts with mismatched Cytosine residues Osmium tetroxide reacts with mismatched Thymidine residues PCR using several primer pairs SIMULTANEOUSLY is known as__________ Multiplex PCR (Colony PCR) To release the DNA from the cells, The PCR is either started with an extended time ___________ 95 C Hot-start PCR is also known as ______. cold-finish PCR Hot-start/cold-finish PCR is achieved with new hybrid polymerases that are inactive at ____________temperature. ambient Hot-start/cold-finish PCR is achieved with new hybrid polymerases that are instantly activated at ___________temperature. elongation What are the 3 main steps included in the process of PCR: 1. Heat Denaturation 2. Primer Annealing 3. Primer Extension What are the Good Primers Characteristic

A melting temperature (Tm) in the range of 52 oC to 65 oC Absence of dimerization capability Absence of significant hairpin formation (>3 bp) PCR was invented in ___________by ____________________. 1984 Kary mullis Extension or polymerization is done by a temprature resistance polymerase named ___________which is extracted from________. Taq p Thermus aquaticus. olymerase Advantages Of PCR? * Rapid cycling (30 minutes to 2 hours). * Specific & sensitive. * Not much more expensive. Disadvantages Of PCR ? * Pollution. * Poor precision. * Hard to get quantitative data. Real time PCR is the most accurate method to detect What? Copy number of each gene Amount of gene expression Efficiency of drugs Virus infection What is SSCP ? Single Strand Conformation Polymerisation Allele Specific Oligomer Hybridisation is also known as ___________. Dot Blot Method A modified form of DGGE is called as_________. TGGE (Temperature gradient gel electrophoresis) Oligonucleotide ligation assay is a process for detection of known______________-, Single Nucleotide Polymorphism OLA is based on joining of 2 oligonucleotide probes i.e, _________ and _________ oligos using_____________. Capture , Reporter , DNA Ligase In CCM mispaired nucleotides are modified by chemical agent using __________, Maxam Gilbert Sequencing PPT is based on combination of what? PCR, Transcription and Translation. Why do 2 probes differ only in sequence at last base at 3end in OLA? To detect the single SNP of known type.

Who invented PCR and when? Kary Mullis, In 1985 What are the three basic steps in PCR technique? Denaturation Primer annealing Extension and DNA Synthesis From which organism DNA polymerase I enzyme is extracted? Thermus aquaticus Why Taq polymerase is extracted from Thermus aquaticus? Because it is thermostable. What nucleotide sequence does taq DNA polymerase add at the end of each strand it synthesises? Adenosine Denaturation temperature and its duration? 94 C for 2 min. Which type of PCR is used to amplify, isolate or identify a known sequence from a cellular or tissue RNA? RT PCR Which type of PCR is used to measure the quantity of a PCR product? Quantitative PCR RACE stands for? Rapid Amplifiacation of cDNA Ends OLA stands for? Oligo Ligation Assay DGGE stands for? Denaturating Gradient Gel Electrophoresis Name a disease which can be diagnosed bt Real Time PCR? HIV Cystic Fibrosis Can bacterial colonies be screened by PCR? YES Which PCR is used to screen bacterial colonies? Colony PCR Xn=X0(1+E)n , Xn stands for? PCR product after cycle n Internal structures which should be avoided in a good primer? Hairpin Self Dimer Dimer What is the unique character of multiplex PCR? It has multiple primers SSCP stands for? Strand Conformation Polymorphism

Which instrument provides provides a simple means of analyzing real-time PCR data from raw fluorescence data? DART-PCR What are the two types of Dyes used in Taqman technique? Reporter dye Quencher dye PCR is discovered by: Kary Mulus At which step primer is attached to fragment Annealing PCR is based on which effect? Peltier effect In PCR the temperature of upper lid is: 120C Why the temperature of upper lid is 120C To prevent the evaporation of cocktail Quantative anaysis of PCR product is done by qRT PCR Full form of FRET: Frequency resonance energy transfer Taqmann assay is based on FRET phenomenon Role of Quencher in taqmann assay: To absorb the fluroscence of probe Clones are identify by hybridizing them with: Probe qRT PCR is mainly used in Disease detection Dye used in qRT PCR SYBR Green SYBR Green Dye binds to_____ groove of DNA minor Annealing temperature is determined by: Tm - 4C Melting temperature is determined by: 4*(G+C)+2*(A+T) SSCP stands for Single stranded conformation polymorphism ASO is stand for Allele specific oligomer hybridization Taq polymerase is isolated from Thermus aquaticus Substitutes of taq polymerase is: PFU and VENT Why E.coli polymerase is not used in PCR reaction It is inactivated or degraded at 94C because E.coli also killed at above 60C QUIZ III By : Randhir Kumar M.Tech. Biotech 1st year CCM is stands for Chemical cleavage methods Primers used in Nested PCR are:

2 Tm of longer than 13 ntds are: Tm= 64.9 +41*(yG+zC-16.4)/ (wA+xT+yG+zC) GC composition in Primer is 50-60% GCG is used for Primer design For amplifying multiple sites ________ PCR is used multiplex Capture and Reporter Oligos are used in Oligonucleotide ligation assay Which type of DNA is known as ancient DNA A DNA Use of DMSO in PCR master mix Decrease secondary structure Why is the PCR cycle doesnt run above 40 cycles? Extremely high chance of non-specific products Molecular beacon is based on the structure? Hairpin loop H1N1 virus strain is detected by: qRT PCR Size of HIV proviral genome 9400 bp PTT is based on PCR, Transcription and Translation Fragment size used in PTT is : 4-5 kb RACE stands for: Rapid amplification of cDNA ends 16s Gene analysis is used for To find out the evolution of the organism Universal primers are used in which technique? 16s gene PCR PCR equation used in qRT PCR is: Xn = X0(1 + E)n DART PCR used for Analyzing real time PCR data Expand FISH Fluorescent in situ hybridisation Which PCR is used to detect phenyl ketonuria? Real time PCR Time duration for Annealing of primer is? 45 sec Time period required for denaturation is 1 min Time duration required for extension is 2 min Formula to calculate melting temp Tm=(wA+xT)*2+(yG+zC)*4 Appropriate GC content in a good primer is

50-60% Expand ASA Allele specific amplification Expand DGGE Denaturing gradient gel electrophoresis Give a variant of DGGE TGGE-temperature gradient gel electrophoresis Give a synonym for hot start Cold finish PCR Which enzyme is used for the joining of capture and reporter oligos DNA ligase Which chemical is used for cleaving DNA:DNA and DNA:RNA hetroduplex in CCM piperidine Use of OLA Detection of SNP Type of folding that takes place in SSCP Intra folding Which method is used for joining of two adjacent oligonucleotide probes? OLA Which type of mutations are detected by PTT? Frameshift and point mutations Allele discrimination occurs by CCM Which technique is used to detect the size of translation products in PTT? Gel electrophoresis/SDS-PAGE Fragment length of DNA sequence analysed by CCM 2 kb Which PCR reduces non-specific amplification during the initial set up stages of the PCR ? HOT START PCR In which PCR, Special chimeric DNA polymerase is used ? Colony PCR Which PCR is used for the Detection of Specific RNAs ? RT-PCR Who discovered PCR ? Kary mullis REST stands for ? Relative expression software tool Which PCR is used in Detection of unknown deleted genes in Thalassemia ? Real Time PCR Give the example of Y-family DNA polymerase ? DNA Poly iv and v

The temperature at which half the DNA strands are single stranded and half are double-stranded is known as ? Melting Temperature The temperature at which primers anneal to the template DNA is called ? Annealing temperature In which PCR Melting temperatures of primer should be similar ? Multiplex PCR Full form of ARMS PCR ? Amplification refractive mutation system PCR Primers which are designed to amplify multiple products ? Universal primers DART-PCR stands for ? Data Analysis for Real-Time PCR What is the relationship b/w fluorescence from reporter dye to the number of amplicons generated in Taqman Assay? Directly proportional PTT is based on a combination of ? PCR, Transcription and Translation. Which test selectively detects translationtermination mutations? PTT (Protein Translation Test) CFLP Stands for.? Cleavage Fragments Length Polymorphism Specific method used for the detection of known SNPs. Oligonucleotide ligation assay Which two types of probes are used in OLA ? Capture and Reporter Oligos Which probes are fluorescently labeled in OLA- Capture or Reporter Oligos? Capture probes Variations in the oligodirected mutagenesis 1.Multiple point 2.Insertion 3.deletion Application of site directed mutagenesis Protein engineering Technique used when gene is present between two restriction enzyme cleavage site Casettee mutagenesis PCR machine is also known as thermocycler Overlapping extensions occurs in SOEing In which technique non-specific amplification reduced during the initial set up stages of the PCR. HOT START PCR transgenic plant screen is done in

Multiplex PCR Special chimeric DNA polymerase are used in Colony PCR SSCP Single-Stranded Conformation Polymophism The temperature at which half the DNA strands are single stranded and half are double-stranded Melting temperature Annealing Temperature Tm_primer 4C Two types of primers Forward Reverse Multiple primers are added in same tube in? Multiplex PCR Real time PCR used for Diagnostic purposes Give one example of polymerase used in PCR Vent polymerase Pfu polymerase Taq DNA polymerase OsO4 mispaired with Thymidine T-vector is? Linear vector For larger fragments than 150 to 200 bp acceptable sensitivities may be achieved, in SCCP RNA-SSCP. 3 to 5 Exonuclease Enzymes are called Proof Readers Name a software for primer designing Primer 3 True or false Tanneal = Tm_primer 4 C True In which PCR Multiple primer pairs can be added in the same tube to do the PCR Multiplex PCR Give one application of Multiplex PCR genome identification ESTIMATE THE NO. OF SEQUENCES IN KCYCLES 2K GIVE ANY TWO CHARACTERISTIC FOR PRIMER DESIGNING? UNIQUNESS

LENGTH BASE COMPOSITION Tm ANNEALING TEMPERATURE INTERNAL STRUCTURE PRIMER PAIR MATCHING ON WHICH NUCLEOTIDE PAIR DO MELTING TEMPERATURE DEPEND? G+C PCR was invented in 1984 by Kary mullis Give three basic steps which are in common with all type of PCR Thermal denaturation Primer annealing Extension or polymerization Name an Excel-based tool for the management and automatic analysis of realtime quantitative PCR data. QBASE Expand DART-PCR Data Analysis for Real-Time PCR Most accurate method to detect Methylation of DNA is Real time PCR Bacterial colonies are screened directly by which PCR COLONY PCR Expand RACE Rapid Amplification of cDNA Ends Random mutation used in which type of detection technique? Single Strand Conformation Polymorphism (SSCP) Name Y-family DNA polymerases? DNA polymerase IV DNA polymerase V 15. Paraffin embedded procedure for DNA extraction used in which mutation technique? Denaturing Gradient Gel Electrophoresis (DGGE) Full form of FASTA? FAST All FULL FORM OF BLAST BASIC LOCAL ALIGNMENT SEARCH TOOL FORMULA FOR ANNEALING TEMPERATURE T(anneal)= Tm-4C

Estimate the No. of sequences that have No. of K-cycles 2K /2 ESTIMATE THE NO. OF SEQUENCES IN KCYCLES 2K GIVE ANY TWO CHARACTERISTIC FOR PRIMER DESIGNING? UNIQUNESS LENGTH BASE COMPOSITION Tm ANNEALING TEMPERATURE INTERNAL STRUCTURE PRIMER PAIR MATCHING ON WHICH NUCLEOTIDE PAIR DO MELTING TEMPERATURE DEPEND? G+C DIFFERENCE BETWEEN ANNEALIN TEMPERATURE AND MELTING TEMPERATURE FOR PRIMER DESIGNING IS_____ 3C GIVE MELTING TEMPERATURE FOR A SEQUENCE SHORTER THAN 13 bp Tm= (wA+xT) * 2 + (yG+zC) * 4 WHICH VIRUS IS USED FOR RT PCR? AMV (AVIAN MOSAIC VIRUS ) WHICH PCR INVOLVES PCR SYNTHESIS? RT PCR WHICH PCR HAS ITS MAJOR APPLICATION IN DIAGNOSTIC PURPOSE? qRT PCR Full form of REST Relative expression software tool Give the PCR equation Xn= Xo (1+E)n Full form of DART PCR Data analysis for real time PCR Which PCR does not require gel based analysis? Real time PCR Give an example of hybridization probe technique used for detection in real time PCR? Light cycler Give an example of hydrolysis probe technique used for detection in real time PCR? Taqman , molecular beacon, Which dye binds with minor grove of DNA? SYBR green SYBER green always binds with double stranded DNA . True or false? True

In dissociation curve, what is the relation between fluorescence and temperature? Inversely proportional How does dissociation curve depicts dimer /falsepriming/contamination It gives a peak To which end does fluorescein binds in a DNA sequence 3 end To which end does Rhodamine binds in a DNA sequence 5 end Which software is used to visualize the probes? FRET WHICH SEQUENCE IS USED BY MOLECULAR BEACON TO DETECT THE NON SPECIFIC INTERACTIONS? HAIRPIN LOOP IN WHICH DISEASE PHENYL ALANINE DEPOSITION OCCURS Phenylketonuria Internal and external primers are the characteristic of which PCR? Nested PCR Which PCR is used for the screening of transgenic plants? Multiplex PCR In PCR script which restriction enzyme is used along with ligase for blunt end coloning Srf 1 Lowering of annealing temperature is the characteristic of which PCR Touchdown PCR In touchdown PCR give the temperature range to which the temperature is lowered in every step 0.5- 1C Colony PCR is used for prokaryotes only true or false? True which PCR use special chimeric sequence? Colony PCR Which technique uses overlapping extension SoEing Which two primers are used in SoEing Internal primers and flanking primers Do insertion of new fragment by SoEing require host ? No Full form of SSCP Single strand conformation polymorphism Example of hybridization based detection SSCP, ASO, melt curve Example of sequencing based detection Sequence specific PCR

Full form of ASO? Allele specific oligomer hybrization Silver staining is used for ____ Low molecular weight protein Example of clevage based detection RFLP Which gel electrophoresis is used for separation of single stranded conformers PAGE AND CGE Protein Translation Test detects mution in___ PCR, Translation & Transcription Which Gel Electrophoresis is used in PTT??? SDS-PAGE PTT detection which mutations? Point & Frameshift mutation In chemical cleavege method, hydroxylamine reacts with___ Cytosine In chemical cleavege method, Osmium tetroxide reacts with___ Thymine In chemical cleavege method, DNA DNA or DNA RNA duplexes are cleaved with___ Piperidine Cloning of PCR Products can be done by___ T-vectors Blunt end Cloning Sticky end Cloning Polymerase IV & V belongs to which family of DNA Polymerase? Y-Family of DNA Polymerase Disease related to protein can be detected by___ ELISA Western Blotting Give another name for DGGE TGGE TGGE stands for ____ Temperature Gradient Gel Electrophoresis Fragment size used in DGGE 1000 bps Name any probe used in OLA. Capture & Reporter Probe In which technique results are analyzed by ABI Gene Maper Software? OLA (Oligonucleotide Ligation Assay) Name the probe used in OLA that differs at 3 end at the last base. Capture Probe CCM is stands for Chemical cleavage methods Primers used in Nested PCR are: 2 Tm of longer than 13 ntds are: Tm= 64.9 +41*(yG+zC-16.4)/ (wA+xT+yG+zC)

GC composition in Primer is 50-60% GCG is used for Primer design For amplifying multiple sites ________ PCR is used multiplex Capture and Reporter Oligos are used in Oligonucleotide ligation assay Which type of DNA is known as ancient DNA A DNA Use of DMSO in PCR master mix Decrease secondary structure Why is the PCR cycle doesnt run above 40 cycles? Extremely high chance of non-specific products Molecular beacon is based on the structure? Hairpin loop H1N1 virus strain is detected by: qRT PCR Size of HIV proviral genome 9400 bp PTT is based on PCR, Transcription and Translation Fragment size used in PTT is : 4-5 kb RACE stands for: Rapid amplification of cDNA ends 16s Gene analysis is used for To find out the evolution of the organism Universal primers are used in which technique? 16s gene PCR PCR equation used in qRT PCR is: Xn = X0(1 + E)n DART PCR used for Analyzing real time PCR data Q. Estimate the total number of sequences after k cycles. 2k Q. Estimate the number of sequences that have unwanted tails after k cycles. 2 A Good Primer has melting temperature (Tm) in the range of 52 oC to 65 oC Absence of dimerization capability is characteristic of a Good Primer Generally speaking, the length of primer has to be at least 15 bases

Usually, average (G+C) content around _______will give us the right melting/annealing temperature for ordinary PCR reactions 50-60% True or false Tanneal = Tm_primer 4C True Name a software for primer designing Primer 3 In which PCR Multiple primer pairs can be added in the same tube to do the PCR Multiplex PCR Give one application of Multiplex PCR genome identification PCR was invented in 1984 by Kary mullis Give three basic steps which are in common with all type of PCR Thermal denaturation Primer annealing Extension or polymerization Name an Excel-based tool for the management and automatic analysis of realtime quantitative PCR data. QBASE Expand DART-PCR Data Analysis for Real-Time PCR Most accurate method to detect Methylation of DNA is Real time PCR Bacterial colonies are screened directly by which PCR COLONY PCR Expand RACE Rapid Amplification of cDNA Ends Dna pol from y-family Pol-IV Pol-V Q.Expand y-family Yeast family ABI gene mapper is used in ? ola