This document describes various types of PCR including inverse PCR, colony PCR, hot start PCR, multiplex PCR, in situ PCR, long PCR, nested PCR, touchdown PCR and their applications. Inverse PCR is used to amplify unknown flanking sequences. Colony PCR screens bacterial colonies without isolating DNA. Hot start PCR prevents nonspecific amplification. Multiplex PCR amplifies multiple targets simultaneously. Nested PCR increases specificity with two primer sets. Touchdown PCR optimizes annealing temperatures.
A real-time polymerase chain reaction is a laboratory technique of molecular biology based on the polymerase chain reaction (PCR). It monitors the amplification of a targeted DNA molecule during the PCR, i.e. in real-time, and not at its end, as in conventional PCR.
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SNP (Single Nucleotide Polymorphic), SNP mapping, SNP profile, SNP types, SNP analysis by gel electropherosis and by mass spectrometry, SNP effects, single strand conformation polymorphism, SNP advantages and disadvantages and application of SNP profile in drug choice
A real-time polymerase chain reaction is a laboratory technique of molecular biology based on the polymerase chain reaction (PCR). It monitors the amplification of a targeted DNA molecule during the PCR, i.e. in real-time, and not at its end, as in conventional PCR.
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SNP (Single Nucleotide Polymorphic), SNP mapping, SNP profile, SNP types, SNP analysis by gel electropherosis and by mass spectrometry, SNP effects, single strand conformation polymorphism, SNP advantages and disadvantages and application of SNP profile in drug choice
Techniques based on the principle of selectively amplifying a subset of restriction fragments from a complex mixture of DNA fragments obtained after digestion of genomic DNA with restriction endonucleases.
complete Single Nucleotide Polymorphiitsm Detection methods with Advance techniques with its applications
Single nucleotide polymorphisms are single base variations between genomes within a species.
There are at least 10 million polymorphic sites in the human genome.
SNPs can distinguish individuals from one another
Denaturing Gradient Gel Electrophoresis
Chemical Cleavage Of Mismatch
Single-stranded Conformation Polymorphism (SSCP)
MutS Protein-binding Assays
Mismatch Repair Detection (MRD)
Heteroduplex Analysis (HA)
Denaturing High Performance Liquid Chromatography (DHPLC)
UNG-Mediated T-Sequencing
RNA-Mediated Finger printing with MALDI MS Detection
Sequencing by Hybridization
Direct DNA Sequencing
Single-feature polymorphism (SFP)
Invader probe
Allele-specific oligonucleotide probes
PCR-based methods
Allele specific primers
Sequence Polymorphism-Derived (SPD) markers
Targeting induced local lesions in genomes (TILLinG)
Minisequencing primers
Allele-specific ligation probes
the speed and ease of use, sensitivity, specificity and robustness of PCR has revolutionized molecular biology and made PCR the most useful and powerful technique with great spectrum of research and diagnostic applications.
PCR is a technique which is used to amplify the number of copies of a specific region of DNA, in order to produce enough DNA to be adequately tested.
Cell-free amplification for synthesizing multiple identical copies (billions) of any DNA of interest.
Basic tool for the molecular biologist.
The purpose of a PCR is to make a huge number of copies of a gene. As a result, it now becomes possible to analyze and characterize DNA fragments found in minute quantities in places like a drop of blood at a crime scene or a cell from an extinct dinosaur.
Like Xerox machine for gene copying.
Sanger sequencing is one of the DNA sequencing methods used to identify and determine the sequence (Nucleotide) of DNA .This is an enzymatic method of sequencing developed by Fred Sanger.
RAPD markers are decamer DNA fragments.
RAPD is a type of PCR reaction.
as the name suggest it is a fast method when compared to the traditional PCR medthod.
Techniques based on the principle of selectively amplifying a subset of restriction fragments from a complex mixture of DNA fragments obtained after digestion of genomic DNA with restriction endonucleases.
complete Single Nucleotide Polymorphiitsm Detection methods with Advance techniques with its applications
Single nucleotide polymorphisms are single base variations between genomes within a species.
There are at least 10 million polymorphic sites in the human genome.
SNPs can distinguish individuals from one another
Denaturing Gradient Gel Electrophoresis
Chemical Cleavage Of Mismatch
Single-stranded Conformation Polymorphism (SSCP)
MutS Protein-binding Assays
Mismatch Repair Detection (MRD)
Heteroduplex Analysis (HA)
Denaturing High Performance Liquid Chromatography (DHPLC)
UNG-Mediated T-Sequencing
RNA-Mediated Finger printing with MALDI MS Detection
Sequencing by Hybridization
Direct DNA Sequencing
Single-feature polymorphism (SFP)
Invader probe
Allele-specific oligonucleotide probes
PCR-based methods
Allele specific primers
Sequence Polymorphism-Derived (SPD) markers
Targeting induced local lesions in genomes (TILLinG)
Minisequencing primers
Allele-specific ligation probes
the speed and ease of use, sensitivity, specificity and robustness of PCR has revolutionized molecular biology and made PCR the most useful and powerful technique with great spectrum of research and diagnostic applications.
PCR is a technique which is used to amplify the number of copies of a specific region of DNA, in order to produce enough DNA to be adequately tested.
Cell-free amplification for synthesizing multiple identical copies (billions) of any DNA of interest.
Basic tool for the molecular biologist.
The purpose of a PCR is to make a huge number of copies of a gene. As a result, it now becomes possible to analyze and characterize DNA fragments found in minute quantities in places like a drop of blood at a crime scene or a cell from an extinct dinosaur.
Like Xerox machine for gene copying.
Sanger sequencing is one of the DNA sequencing methods used to identify and determine the sequence (Nucleotide) of DNA .This is an enzymatic method of sequencing developed by Fred Sanger.
RAPD markers are decamer DNA fragments.
RAPD is a type of PCR reaction.
as the name suggest it is a fast method when compared to the traditional PCR medthod.
Event Management System Vb Net Project Report.pdfKamal Acharya
In present era, the scopes of information technology growing with a very fast .We do not see any are untouched from this industry. The scope of information technology has become wider includes: Business and industry. Household Business, Communication, Education, Entertainment, Science, Medicine, Engineering, Distance Learning, Weather Forecasting. Carrier Searching and so on.
My project named “Event Management System” is software that store and maintained all events coordinated in college. It also helpful to print related reports. My project will help to record the events coordinated by faculties with their Name, Event subject, date & details in an efficient & effective ways.
In my system we have to make a system by which a user can record all events coordinated by a particular faculty. In our proposed system some more featured are added which differs it from the existing system such as security.
Saudi Arabia stands as a titan in the global energy landscape, renowned for its abundant oil and gas resources. It's the largest exporter of petroleum and holds some of the world's most significant reserves. Let's delve into the top 10 oil and gas projects shaping Saudi Arabia's energy future in 2024.
About
Indigenized remote control interface card suitable for MAFI system CCR equipment. Compatible for IDM8000 CCR. Backplane mounted serial and TCP/Ethernet communication module for CCR remote access. IDM 8000 CCR remote control on serial and TCP protocol.
• Remote control: Parallel or serial interface.
• Compatible with MAFI CCR system.
• Compatible with IDM8000 CCR.
• Compatible with Backplane mount serial communication.
• Compatible with commercial and Defence aviation CCR system.
• Remote control system for accessing CCR and allied system over serial or TCP.
• Indigenized local Support/presence in India.
• Easy in configuration using DIP switches.
Technical Specifications
Indigenized remote control interface card suitable for MAFI system CCR equipment. Compatible for IDM8000 CCR. Backplane mounted serial and TCP/Ethernet communication module for CCR remote access. IDM 8000 CCR remote control on serial and TCP protocol.
Key Features
Indigenized remote control interface card suitable for MAFI system CCR equipment. Compatible for IDM8000 CCR. Backplane mounted serial and TCP/Ethernet communication module for CCR remote access. IDM 8000 CCR remote control on serial and TCP protocol.
• Remote control: Parallel or serial interface
• Compatible with MAFI CCR system
• Copatiable with IDM8000 CCR
• Compatible with Backplane mount serial communication.
• Compatible with commercial and Defence aviation CCR system.
• Remote control system for accessing CCR and allied system over serial or TCP.
• Indigenized local Support/presence in India.
Application
• Remote control: Parallel or serial interface.
• Compatible with MAFI CCR system.
• Compatible with IDM8000 CCR.
• Compatible with Backplane mount serial communication.
• Compatible with commercial and Defence aviation CCR system.
• Remote control system for accessing CCR and allied system over serial or TCP.
• Indigenized local Support/presence in India.
• Easy in configuration using DIP switches.
Quality defects in TMT Bars, Possible causes and Potential Solutions.PrashantGoswami42
Maintaining high-quality standards in the production of TMT bars is crucial for ensuring structural integrity in construction. Addressing common defects through careful monitoring, standardized processes, and advanced technology can significantly improve the quality of TMT bars. Continuous training and adherence to quality control measures will also play a pivotal role in minimizing these defects.
Courier management system project report.pdfKamal Acharya
It is now-a-days very important for the people to send or receive articles like imported furniture, electronic items, gifts, business goods and the like. People depend vastly on different transport systems which mostly use the manual way of receiving and delivering the articles. There is no way to track the articles till they are received and there is no way to let the customer know what happened in transit, once he booked some articles. In such a situation, we need a system which completely computerizes the cargo activities including time to time tracking of the articles sent. This need is fulfilled by Courier Management System software which is online software for the cargo management people that enables them to receive the goods from a source and send them to a required destination and track their status from time to time.
Explore the innovative world of trenchless pipe repair with our comprehensive guide, "The Benefits and Techniques of Trenchless Pipe Repair." This document delves into the modern methods of repairing underground pipes without the need for extensive excavation, highlighting the numerous advantages and the latest techniques used in the industry.
Learn about the cost savings, reduced environmental impact, and minimal disruption associated with trenchless technology. Discover detailed explanations of popular techniques such as pipe bursting, cured-in-place pipe (CIPP) lining, and directional drilling. Understand how these methods can be applied to various types of infrastructure, from residential plumbing to large-scale municipal systems.
Ideal for homeowners, contractors, engineers, and anyone interested in modern plumbing solutions, this guide provides valuable insights into why trenchless pipe repair is becoming the preferred choice for pipe rehabilitation. Stay informed about the latest advancements and best practices in the field.
COLLEGE BUS MANAGEMENT SYSTEM PROJECT REPORT.pdfKamal Acharya
The College Bus Management system is completely developed by Visual Basic .NET Version. The application is connect with most secured database language MS SQL Server. The application is develop by using best combination of front-end and back-end languages. The application is totally design like flat user interface. This flat user interface is more attractive user interface in 2017. The application is gives more important to the system functionality. The application is to manage the student’s details, driver’s details, bus details, bus route details, bus fees details and more. The application has only one unit for admin. The admin can manage the entire application. The admin can login into the application by using username and password of the admin. The application is develop for big and small colleges. It is more user friendly for non-computer person. Even they can easily learn how to manage the application within hours. The application is more secure by the admin. The system will give an effective output for the VB.Net and SQL Server given as input to the system. The compiled java program given as input to the system, after scanning the program will generate different reports. The application generates the report for users. The admin can view and download the report of the data. The application deliver the excel format reports. Because, excel formatted reports is very easy to understand the income and expense of the college bus. This application is mainly develop for windows operating system users. In 2017, 73% of people enterprises are using windows operating system. So the application will easily install for all the windows operating system users. The application-developed size is very low. The application consumes very low space in disk. Therefore, the user can allocate very minimum local disk space for this application.
CFD Simulation of By-pass Flow in a HRSG module by R&R Consult.pptxR&R Consult
CFD analysis is incredibly effective at solving mysteries and improving the performance of complex systems!
Here's a great example: At a large natural gas-fired power plant, where they use waste heat to generate steam and energy, they were puzzled that their boiler wasn't producing as much steam as expected.
R&R and Tetra Engineering Group Inc. were asked to solve the issue with reduced steam production.
An inspection had shown that a significant amount of hot flue gas was bypassing the boiler tubes, where the heat was supposed to be transferred.
R&R Consult conducted a CFD analysis, which revealed that 6.3% of the flue gas was bypassing the boiler tubes without transferring heat. The analysis also showed that the flue gas was instead being directed along the sides of the boiler and between the modules that were supposed to capture the heat. This was the cause of the reduced performance.
Based on our results, Tetra Engineering installed covering plates to reduce the bypass flow. This improved the boiler's performance and increased electricity production.
It is always satisfying when we can help solve complex challenges like this. Do your systems also need a check-up or optimization? Give us a call!
Work done in cooperation with James Malloy and David Moelling from Tetra Engineering.
More examples of our work https://www.r-r-consult.dk/en/cases-en/
Automobile Management System Project Report.pdfKamal Acharya
The proposed project is developed to manage the automobile in the automobile dealer company. The main module in this project is login, automobile management, customer management, sales, complaints and reports. The first module is the login. The automobile showroom owner should login to the project for usage. The username and password are verified and if it is correct, next form opens. If the username and password are not correct, it shows the error message.
When a customer search for a automobile, if the automobile is available, they will be taken to a page that shows the details of the automobile including automobile name, automobile ID, quantity, price etc. “Automobile Management System” is useful for maintaining automobiles, customers effectively and hence helps for establishing good relation between customer and automobile organization. It contains various customized modules for effectively maintaining automobiles and stock information accurately and safely.
When the automobile is sold to the customer, stock will be reduced automatically. When a new purchase is made, stock will be increased automatically. While selecting automobiles for sale, the proposed software will automatically check for total number of available stock of that particular item, if the total stock of that particular item is less than 5, software will notify the user to purchase the particular item.
Also when the user tries to sale items which are not in stock, the system will prompt the user that the stock is not enough. Customers of this system can search for a automobile; can purchase a automobile easily by selecting fast. On the other hand the stock of automobiles can be maintained perfectly by the automobile shop manager overcoming the drawbacks of existing system.
Welcome to WIPAC Monthly the magazine brought to you by the LinkedIn Group Water Industry Process Automation & Control.
In this month's edition, along with this month's industry news to celebrate the 13 years since the group was created we have articles including
A case study of the used of Advanced Process Control at the Wastewater Treatment works at Lleida in Spain
A look back on an article on smart wastewater networks in order to see how the industry has measured up in the interim around the adoption of Digital Transformation in the Water Industry.
Democratizing Fuzzing at Scale by Abhishek Aryaabh.arya
Presented at NUS: Fuzzing and Software Security Summer School 2024
This keynote talks about the democratization of fuzzing at scale, highlighting the collaboration between open source communities, academia, and industry to advance the field of fuzzing. It delves into the history of fuzzing, the development of scalable fuzzing platforms, and the empowerment of community-driven research. The talk will further discuss recent advancements leveraging AI/ML and offer insights into the future evolution of the fuzzing landscape.
3. INVERSE PCR
Inverse PCR is used to amplify unknown DNA that flanks one end of
known DNA sequence for which no primers are available .
In this PCR information of only one internal sequence of the target DNA is
required. So, It is very useful in identifying flanking DNA sequences of
genomic inserts.
This technique involves digestion by restriction enzyme of a DNA
preparation having known DNA sequence and its flanking sequence. The
restriction fragments are self-ligated and changed into circular DNA and the
circularized DNA is used as a template in the PCR reaction.
The primers are designed from known sequence that faces outward from
that region .The resulting amplified product will be a single linear fragment
that includes unknown DNA from both left and right sides. The PCR
product can be cloned and then sequenced.
3
5. Inverse PCR Applications
Amplification and identification of sequences flanking
transposable elements
Identification of genomic inserts
Cloning of unknown cDNA sequences from total RNA
Construction of end-specific probes for chromosome walking
Amplification of integration sites used by viruses and
transgenes.
5
6. COLONY PCR
Colony PCR is used for screening recombinants from
bacterial, bacteriophage or yeast transformation products.
A colony is picked with a sterile toothpick or pipette tip
from a growth plate.
This is then inserted into autoclaved water and PCR is
then conducted
6
7. Applications of colony PCR
Colony PCR is a fast and reliable method for the
screening of recombinants.
A number of colonies or plaques can be assayed
simultaneously and there is no need to store a large
number of transformed clones for long periods.
This method can easily be used for cDNA library
screening.
7
8. HOT START PCR
Hot Start PCR permits the inhibition of polymerase activity during
setting of PCR reaction, by limiting polymerase activity prior to PCR
cycling.
The primers, Mg2++, buffer and dNTPs can be mixed at room
temperature in the bottom of the PCR tube and then covered with
melted wax (e.g., Ampliwax PCR Gems from Perkin-ELMER). The
wax solidifies on cooling and limits the reagents to bottom of the
tube.
The remaining components are then added on top of the barrier. Wax
layer melts upon heating during the denaturation step and the two
aqueous layers get mixed resulting in a fully active reaction.
The melted wax floats to the top of the reaction mixture where it acts
as a barrier to evaporation.
8
9. Automated Hot Start PCR
More recently, an alternate method of automated Hot Start PCR
has been developed where Taq DNA polymerase-directed
monoclonal antibodies, as thermolabile inhibitors of enzymes,
are used.
At low temperature, i.e., at ambient temperature antigen-
antibody interaction results in potent inhibition of DNA
polymerase;
When temperature rises at the start of thermocycling process,
the antibodies are denatured and fully active Taq DNA
polymerase is released.
The pre-incubation of Taq DNA polymerase and antibodies
results in highly specific PCR reaction with increased
sensitivity.
9
10. Applications of Hot Start PCR
Hot Start PCR reduces nonspecific amplification and
increases the sensitivity, specificity, precision of
amplification of low copies of target DNA and yield of
PCR product.
10
11. MULTIPLEX PCR
All sequences of interest are simultaneously amplified in
multiplex PCR.
Multiplex PCR has significant template, time and cost
saving advantages, especially when a large number of
individual sequences need to be analyzed. Single aliquot
of DNA or RNA is required rather than an aliquot for each
marker to be analyzed.
Generally up to eight primer pairs can be used in a
standard multiplex reaction, otherwise the yield of some
amplicons is reduced and not visible on agarose gel.
11
13. Applications of Multiplex PCR
This type of PCRs is important for forensic application,
prenatal diagnosis, and for clinical applications in which
the tissue/DNA samples are limited.
It is also used for genotyping applications where
simultaneous analysis of multiple markers is required.
13
14. In situ PCR (IS-PCR)
It is primer driven amplification of a DNA or RNA
template by PCR and its subsequent detection within the
histological tissue section or cell preparation.
Preparation of sample for IS-PCR, i.e., fixation and
permeabilization
Actual PCR
Detection of amplified signal
It is somewhat difficult to detect the genes of low copy
number by in situ PCR as it is below the detection limit.
14
15. Applications of IS-PCR
In situ PCR amplification can be performed on fixed
tissue or cells or on a slide .
15
16. Long PCR
Long PCR is a PCR, which is extended longer than standard PCR,
over 5 kbp (frequently. over 10 kbp).
In case of standard PCR it is very difficult to get long PCR product
because of damage to the template and product DNAs due to
exposure to high temperature, presence of Mn2+ and difficulties in
denaturing of long DNA molecules.
There are recent reports of amplification of 42 kbp with the blend of
enzymes primarily containing non-proofreading polymerase with a
very small amount of proofreading polmerase.
(i)45:1 Able to amplify a 22 kbp fragment of ß globin
from genomic DNA
(ii)125:1 Able to amplify 39 kbp of λ DNA
16
17. Nested PCR
In case of nested PCR two or more pairs instead of one
pair of PCR primers are used to amplify a fragment.
The first pair of PCR primers amplifies a fragment similar
to a standard PCR.
Internal primers, called nested primers as these hybridize
to the sites nested within the first fragment, are used to
amplify PCR products formed by external primers.
The binding of nested primers inside the first PCR product
fragment allows amplification of a second PCR product
that is shorter than the first one.
17
19. Advantages of Nested PCR
The advantage of nested PCR is that if the wrong PCR
fragment is amplified, the probability is quite low that the
region is amplified a second time by the second set of
primers.
Thus, nested PCR is used to increase magnitude and
specificity of amplification.
19
20. Touchdown PCR
In touchdown PCR, the Ta, during the first two cycles is
set at ~3oC above the calculated Ta.
The annealing temperature is then reduced by one degree
centigrade for every one or two cycles.
When the Ta, reaches at an optimum point, then specific
amplification of the target DNA starts.
The onset of nonspecific amplification is delayed for few
additional cycles until the Ta, is lowered to the point where
nonspecific amplification priming can occur.
20
21. Applications of Touchdown PCR
It is used to optimize yield of amplified product at
different annealing temperatures.
In some cases it is very tricky to amplify DNA because of
mismatches between oligonucleotide primers and the
template DNA and it is not possible to calculate annealing
temperature.
21