sequencing-primer-generator

A python tool that will batch-generate optimal DNA primers for Sanger sequencing. The output .xlsx file is directly compatible for bulk input into IDT Oligo Entry.

Introduction

In this repository, there are two files: Primerwalk_GUI.py and Primerwalk.py. Primerwalk_GUI.py is a front end desktop application that communicates with the backend Primerwalk.py python file. This repository would be most helpful for molecular biologist who frequently conduct full plasmid sequencing.

There are licensed softwares that provide primer synthesis capabilities, but none that can synthesize appropriate forward and reverse primers in batch. The exported files of Primerwalk.py is compatible for single tube ordering and 96-well plate ordering from IDT (idtdna.com).

Single tube ordering form: https://www.idtdna.com/site/order/oligoentry
96-well plate ordering form: https://www.idtdna.com/site/order/plate/index/pico/5647

How to use

Python version and installing required packages

Python version: 3.7.3
Installing packages:
$ pip install -r requirements.txt

Starting application

$ python Primerwalk_GUI.py

Input file formatting

Ensure that Primerwalk_GUI.py, Primerwalk.py, sequence_file.csv are stored in the same directory.

Please adhere precisely to the headers in sequence_file.csv, outlined below (note: the headers are case sensitive - please adhere as exactly as listed):

sequence_file.csv

Name: Name of plasmid / DNA sample
Full Sequence: Full sample DNA sequence (case insensitive)
Primer Walk: Sequence of DNA to be sequenced (this could equate to Full Sequence, if desired - case insensitive)
BP Gap: Base pair gap between primers (usually, max is 500 bp before sequence resolution is compromised)
Primer conc. (nM): Primer concentration to order (usually 25nM or 100nM) - used for single tube ordering

Running Primerwalk_GUI.py on terminal

1. Navigate through your terminal to the directory in which Primerwalk_GUI.py is installed.
   
2. Start Primerwalk_GUI.py application
   
3. The following desktop application should appear:
   

Choosing Single Tubes:

If you would like to order single tubes of primers from IDT, click the 'SINGLE TUBES' button on the desktop application. When executed, the desktop application would look like this:

A new directory with today's date will be created in your main directory (i.e. the directory with your Primerwalk_GUI.py, Primerwalk.py, and sequence_file.csv files).
Inside the directory, you will find unique .xlsx files for every plasmid you inputted in the 'sequence_file.csv' file.

Each file will have the following naming format: IDT_Tube_(plasmid name).xlsx
The .xlsx file will look like this:

Upload this file directly to this website and perform the following tasks:

Click on button 'Bulk Input' in center of webpage (blue button)



Click on button 'Choose File' to upload your .xlsx ordering forms



All required fields in the ordering form should automatically fill. Proceed to checkout if satisfied with input.

Choosing 96-Well Plate:

If you would like to order your primers in a 96-well plate, click the '96-WELL PLATE' button on the desktop application. When executed, the desktop application will look like this:

Similar to the output file generated when selecting single tubes, a new directory with the name of today's date will be created in your main directory (i.e. the directory with your Primerwalk_GUI.py, Primerwalk.py, and sequence_file.csv files).
Inside the directory, you will find unique .xlsx files for every plasmid you inputted in the 'sequence_file.csv' file.

Each file will have the following naming format: IDT_Plate_(plasmid name).xlsx

Upload this file directly to this website and perform the following tasks:

Click on button 'Upload Plate(s)' in center of webpage (blue button)



Click on button 'Choose File' to upload your .xlsx ordering forms



All required fields in the ordering form should automatically fill. Proceed to checkout if satisfied with input.

Fin