Agenda • Background • Next Generation Sequencing • Sequence Assembly • Sequence Alignment • Traditional Alignment Algorithms • Next Generation Alignment Algorithms • Conclusion Andy Nagar 2 Background • Earlier sequencing methods were based on Sanger sequencing, which goes back to the 1970s. • Sequencing was slow, bases were read one at a time. • Separation is done by electrophoresis. • Readout by fluorescent tags ...
Our Mtb DNA on the Tapestation MinION fastq file 2 records – 576 bp; 1316 bp Illumina fastq file all records ~100 bp There are 3 main approaches to sequencing Market Approach / USP Read Achievements Ideal for: dominance length st 1 Generation Applied Sequencing by 1 kb Most primary PCR products (Sanger) Biosystems synthesis (SBS) drafts, e.g. (genotyping) - Low throughput Mtb, human ...
Courtesy of CRC Press/Taylor & Francis Group Figure 11.2 Sanger sequencing with fluorescent nucleotides. Rather than using four separate radioactive nucleotides in the sequencing reactions, use of nucleotides each labeled with a different type of fluorescence reduced both the number of reactions (top) and the number of lanes on the polyacrylamide gel (middle). Fluorescence also enabled automated detection (bottom), reducing human error in ...
Objectives Microbial genomes Common isolate identification techniques using molecular biology Whole genome sequencing (WGS) Example of WGS for outbreak investigation Questions Microbial Genomes Genome size varies from 4.56 to 5.70 Mb How big is 5 Mb??? Harry Potter Story Long story, some big books! 1,084,440 words in all seven books Average word length ~5 letters ~5,422,200 letters total in ...
DNA fragment sequencing In order to establish exactly what changes have occurred in the gene under study, it will be necessary determine the DNA sequence of that gene. This is now a routine procedure in molecular biology laboratories, extending not just to individual genes or fragments but to the complete sequence of the DNA (the genome) of the organism. The two main in vitro DNA ...
Next-Generation Sequencing y r o t a r o b ay Lt i ss er ce nv ei in cU S lg an nu oo iY t a tm ua ph mg oi r CB Problem Statement y r o t a r o •Map next-generation sequence reads b a y L t i with variable nucleotide confidence to s s e r c ...
To review before and/or after this session • National record of discussion forms (NHSE/I and Genomics England): oAdults with capacity, parents of children and for deceased individuals oParticipation in the NGRL* oAssent for young persons* oConsultee for adults without capacity* oWithdrawal form* *NGRL only Annotated versions are available for all forms with guide for completion • Whole genome sequencing test order forms: oRare disease proband ...
Why? We want to understand and control transmission jdhd of infectious diseases in Africa to save lives and protect an already fragile health system How? By establishing a continental consortium of key national laboratories, academic and public health institutions to advance outbreak surveillance and research in Africa What? Generate and analyze the genomic diversity of Pathogens in real-time to map the spread of the disease ...
Need to Add to part 2 or 3 A. Chip-seq B. Deep sequencing for expression profiling C. Illumina? movie Genomics Summary A. Microarrays: expression profiling and other uses B. Global Gene Knockouts C. Global protein localization in yeast D. Global complex identification in yeast E. Global two-hybrid analysis in yeast and other organisms F. RNAi G. Transgenics, gene “knock-outs” (genetics not genomics) H. Human ...
Types of Expectation • Nature of samples Human Male Liver • Nature of data RNA-Seq/Genomic • Efficacy of processing Equal losses • Effect of interventionsDid they work • Nature of effects Global/Local • Sources of variation Any unexpected Raw Data Expectation Raw Data Expectations • Bisulphite Sequencing –Whole genome – all regions equally sampled –Both strands – no read level strand bias RNA-Contamination ...
Content Forensic Science and DNA fingerprinting Invention of DNA fingerprinting Preparation of DNA Fingerprint Steps of DNA fingerprinting Types of Analysis RFLP, PCR, STR Analysis, AFLP Analysis Applications DNA Fingerprinting and Forensic Science Forensic science Intersection of Law and Science Historic examples of collecting evidences: 18th Century – Photography Obsolete & Can Early 19th Century – Fingerprints Be Manipulated From 1985 – DNA ...
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