Overview pdf summarising all 8 steps

Getting Started - Overview pdf

STEP 1: Extracting Daffodil Chloroplast DNA

Extracting chloroplasts using ultracentrifugation protocol with buffer instructions pdf

STEP 2. Choosing the Best DNA

Using Gel Electrophoresis to selecting the cleanest DNA sample

For more information visit the mini pcr website at https://www.minipcr.com/products/bluegel/

To program the thermocycler, download the software at https://www.minipcr.com/downloads/

STEP 3: DNA Library Preparation

Walkthrough Video 1: DNA Extraction using Qiagen DNeasy Plant Mini Kit

Extract DNA Qiagen Instructions pdf

STEP 4: DNA Sequencing using MinION Oxford Nanopore Technology

Walkthrough Video 2: DNA Library Preparation Minion Flow Cell Loading & Set-up

Lambda quality assurance protocol pdf

Library prep and flow cell loading instructions pdf

STEP 5: Base Calling using Guppy

Base calling is the process of translating the electronic raw signal of the sequencer into bases. Guppy contains the Oxford Nanopore Technologies’ basecalling algorithms and several bioinformatic post-processing features such as filtering of low quality reads.

Walkthrough Video 3: Base Calling Using Guppy

The code used in the video is available in the description section of the YouTube video (https://youtu.be/PvKkEtChPB0)

Code used in the video:
"C:\Program Files\OxfordNanopore\ont-guppy-cpu\bin\guppy_basecaller.exe" --input_path C:\Users\Science\Documents\30MAR22FURNESS\30mar22furness\30mar22furness\20220330_1607_MN32710_FLG001_9a1e9e66\fast5 --save_path C:\Users\Science\Documents\30MAR22FURNESS\30mar22furness\30mar22furness\30MAR22Furness --flowcell FLO-FLG001 --kit SQK-RAD004

However if you are importing the files into a folder such as "test", you would need to locate this folder within the directory (eg Users\Science\Documents may need to change):
"C:\Program Files\OxfordNanopore\ont-guppy-cpu\bin\guppy_basecaller.exe" --input_path C:\Users\Science\Documents\test --save_path C:\Users\Science\Documents\testbasecalled --flowcell FLO-FLG001 --kit SQK-RAD004

STEP 6: Assembling the DNA sequence with Geneious Prime

Walkthrough Video 4: Geneious Prime Sequence Assembly Instructions

STEP 7: Viewing and Analysising the DNA Sequence in Jalview

Jalview is one of the most widely used programs for visualizing and analyzing DNA, RNA and protein multiple sequence alignments. It uses an interactive multi-window interface for viewing sequences, trees, and three-dimensional structures. It has a number of analysis tools for aligning sequences, measuring similarities and comparing structures.