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===== illumina Sequencing Run 1 ===== ==== Meta ==== Prepared by Akram Tariq Sial Date: Overview: This sample was prepared according to manufacturer's recommendations. [Add link to illumina protocol] ==== Starting DNA ==== ==== Initial Size Selection ==== {{:lab_protocols:bana_slug_library_illumina.jpg?600}} {{:lab_protocols:bana_slug_library_illumina._after_cutjpg.jpg.jpg?600}} [[http://www.fermentas.com/en/products/all/dna-electrophoresis/generuler-dna-ladders/sm0373-generuler-50bp|Fermentas O'generuler 50bp ladder used in these gels.]] Lanes 1 & 6 1µg of 98.0 ng/µl genomic DNA was sheared using either Covaris or a Nebulizer. Lanes 3&4: Banana Slug DNA sheared using Covaris. Lanes 5&6: Banana Slug DNA sheared using a Nebulizer. Both were cut between 250-375 basepairs. ==== Bioanalyzer results of initial shearing and final amplified library size selection ==== {{:lab_protocols:bioanalyzer-illumina-library.pdf|Bioanalyzer results of shearing distributions and final amplified libraries.}} Covaris and Nebulizer shearing are in lanes 1 and 2 respectively. Amplified and size selected libraries for Nebulizer and Covaris in lanes 3 and 4 respectively. The results indicate the final nebulizer library ranges from 202-320bp in distribution. The final covaris library ranges from 225-263bp in length. ==== Final distribution of paired end reads ==== Each stage indicates a different distribution in the length of the paired-end reads. The original size selection cut indicates a fragment size of 250-375. However, the bioanalyzer results indicate the final size selected fragment is much shorter at 200-320 bp for a final distribution between 100-230 bp. ==== Differences between lanes in illumina run ==== Each lane contained a different concentration of starting material used in bridge amplification. ^Lane ^ Starting amount of library and library type ^ |Lane 1 | 8 picoMoles Nebulized Library | |Lane 2 | 10 picoMoles Nebulized Library | |Lane 3 | 12 picoMoles Nebulized Library | |Lane 4 | 6 picoMoles phix control* | |Lane 5 | 8 picoMoles Covaris Library | |Lane 6 | 10 picoMoles Covaris Library | |Lane 7 | 12 picoMoles Covaris Library | |Lane 8 | 14 picoMoles Covaris Library | These different starting quantities of DNA are reflected in the number of clusters generated and the total reads outputted for each lane. Additionally, a different distribution of distance for the paired-end reads are different for lanes 1-3 and 5-8 *Data from lane 4 of the illumina run **SHOULD NOT** be used for assembly
