Banana Slug Genomics

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Lecture Notes 4/6/2015

Note Taker: Christopher Kan

A road map to the Denovo-Assembly of the Banana Slug Genome
	- Stefan Prost

Denovo VS. Reference Genome
	- Reference can be biased by the assembly itself. Eg some areas may not be annotated or reads are not available. 
	- Denovo costs more

Scaffolds and Contigs
	- Contigs have little to no gaps
	- Scaffolds can have missing regions but the linear order of the contigs within each scaffold is known
	- N50s for Scaffold and Contigs are used as quality measures. 
		○ You sum the size of the scaffolds or contigs until you reach 1/2 the linear length of a genome. The size of the last constituent part of the N50. It’s a way to obtain a median-esque measure of assembly quality
	- Ideally # scaffolds = # chromosomes

Definition: Kmer - Short unique element of DNA of a certain length n
	- The elements can overlap
	- Used to summarize data by assemblers

A priori knowledge of a genome
	- Expected Genome Size
		* C-values from www.genomesize.com
		* C-value is the genome size in picrograms
		* 1pg=1C=980MB
		* Depending on clade information from related genomes can be used to provide a-priori knowledge
			§ Some have low variation and high synteny - Birds
		* 6-7 GB becomes difficult
	- Data bases
		* www.Gigaadb.org
		* NCBI Genome
	- Expected repeat content
		* Correlated with genome size
		* Small repeats and pseudogenes, genome duplications
	- Expected Heterozygosity
	- Haploid? Diploid or polyploid?
		* No assembler that can assemble polyploid currently
	
Sequencing Technology
	- 1st Gen
		○ Sanger
	- 2nd Gen (PCR Needed)
		* Illumia
			§ It took me a long time to understand how this works these two video helped me: [[https://www.youtube.com/playlist?list=PLfvYDg0hWvoqfF9z7bw7Zizeenj620r5c| Link]]
		* Roche:454
		* IONtorrent
		* ABI: Solid
	- 3rd Gen (Single Molecule Sequencing)
		* Heliscope
		* PacBio RS II
			§ Problems
				* Polyerase needs to be fast with low error
				* Poor yield from cell
					~ Need to wash with low concentration to ensure most cells only have one molocule
				* Insertions and deletions. Missing or having one that hangs around
					~ Random. This property used to error correct
			* Light emission at time of amplification
				* Real time, allows decernment of 3D structure of molocule based on time between incorporations
			* Can circularize small DNA fragments and get multiple reads ~3kb, possible to 8kb
				
		* MiniION and GridION
			* Sequences by taking molocule apart
			* Nanopore allows the molocule through based on salt gradient
				* Sequence as molocule goes through
					~ Molocule held by molocule that clips off one nucleotide at a time - exonuclease
					~ Measure the charge at the nanopore. 
				* OR sequence the molocule as it goes through as its held with an helicase
			* Some systematic errors - Harder to correct 
			* Can use a hair pin to run both side of DNA so its effectiely paired
			* No restriction on size hypothetically
Issues with 3rd Gen
	- High error
	- High cost
	- Error correction very computationally expensive
	




Note Taker: XXX
Banana Slug Genomics

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