Plasmid-ID Kit
(Kit # 1000000168 )

Depositing Lab:   Philip Poole

The Plasmid-ID Kit allows for the high-throughput assessment of competitive and effective rhizobial strains.

This kit will be sent as bacterial glycerol stocks in 96-well plate format.

Original Publication

Optimizing Rhizobium- legume symbioses by simultaneous measurement of rhizobial competitiveness and N ₂ fixation in nodules Mendoza-Suárez MA, Geddes BA, Sánchez-Cañizares C, Ramírez-González RH, Kirchhelle C, Jorrrin B, Poole PS. Proc Natl Acad Sci U S A. 2020 May 5;117(18):9822-9831. doi: 10.1073/pnas.1921225117. Epub 2020 Apr 21. PubMed PMID: 32317381

Description

The Plasmid-ID kit allows the high-throughput assessment of competitive and effective rhizobial strains.

These plasmids are provided in the strain ST18 (S17 delta pir delta hemA), which requires supplementation with 50 μg/mL of ALA (5-aminolevulinic acid) for growth.

The Plasmid-ID help us to:

• Directly observe plant-bacterial interactions
• Assess competitiveness of strains without reducing their fitness
• Obtain information about their nitrogen fixation rates
• Identify rhizobia directly from individual nodules

This plasmid library has the potential to help pre-evaluate strains as possible inoculants on a selected legume crop in local soils before launching large-scale field use.

A.) Plasmid-ID Library built by Golden Gate cloning. The plasmid library includes 95 individual and unique reporter plasmids, with the following modules: vector; pOGG026, PU module; PsnifH from pOGG043, SC module; sfGFP from pOGG037 and the product from annealing the oligonucleotide was added as a T module (Plasmid ID) as the unique identifier in each case. High-throughput conjugation was made possible with the use of chemically competent E. coli ST18 cells for transformations, which allowed a biparental mating. B.) The Plasmid-ID library allows simultaneous high-throughput measurement of N₂ fixation in individual nodules using green fluorescent protein (GFP) and barcode strain identification (Plasmid ID) through next generation sequencing (NGS).

Kit Documentation

Script to identify IDs from sequencing reads available in GitHub: Plasmid-ID

Protocol for high-throughput conjugation from E. coli ST18 into rhizobial strains in 96-well microtiter plates available in the Supplemental Documents of the associated publication

How to Cite this Kit

These plasmids were created by your colleagues. Please acknowledge the Principal Investigator, cite the article in which they were created, and include Addgene in the Materials and Methods of your future publications.

For your Materials and Methods section:

“The Plasmid-ID Kit was a gift from Philip Poole (Addgene kit #1000000168).”

For your Reference section:

Optimizing Rhizobium- legume symbioses by simultaneous measurement of rhizobial competitiveness and N ₂ fixation in nodules Mendoza-Suárez MA, Geddes BA, Sánchez-Cañizares C, Ramírez-González RH, Kirchhelle C, Jorrrin B, Poole PS. Proc Natl Acad Sci U S A. 2020 May 5;117(18):9822-9831. doi: 10.1073/pnas.1921225117. Epub 2020 Apr 21. PubMed PMID: 32317381

Plasmid-ID Kit - #1000000168

Inventory

Well	Plasmid	Resistance
A / 1	pOPS0491	Kanamycin
A / 2	pOPS0492	Kanamycin
A / 3	pOPS0493	Kanamycin
A / 4	pOPS0494	Kanamycin
A / 5	pOPS0495	Kanamycin
A / 6	pOPS0496	Kanamycin
A / 7	pOPS0497	Kanamycin
A / 8	pOPS0498	Kanamycin
A / 9	pOPS0499	Kanamycin
A / 10	pOPS0500	Kanamycin
A / 11	pOPS0501	Kanamycin
A / 12	pOPS0502	Kanamycin
B / 1	pOPS0503	Kanamycin
B / 2	pOPS0504	Kanamycin
B / 3	pOPS0505	Kanamycin
B / 4	pOPS0506	Kanamycin
B / 5	pOPS0507	Kanamycin
B / 6	pOPS0508	Kanamycin
B / 7	pOPS0509	Kanamycin
B / 8	pOPS0510	Kanamycin
B / 9	pOPS0511	Kanamycin
B / 10	pOPS0512	Kanamycin
B / 11	pOPS0513	Kanamycin
B / 12	pOPS0514	Kanamycin
C / 1	pOPS0515	Kanamycin
C / 2	pOPS0516	Kanamycin
C / 3	pOPS0517	Kanamycin
C / 4	pOPS0518	Kanamycin
C / 5	pOPS0519	Kanamycin
C / 6	pOPS0520	Kanamycin
C / 7	pOPS0521	Kanamycin
C / 8	pOPS0522	Kanamycin
C / 9	pOPS0523	Kanamycin
C / 10	pOPS0524	Kanamycin
C / 11	pOPS0525	Kanamycin
C / 12	pOPS0526	Kanamycin
D / 1	pOPS0527	Kanamycin
D / 2	pOPS0528	Kanamycin
D / 3	pOPS0529	Kanamycin
D / 4	pOPS0530	Kanamycin
D / 5	pOPS0531	Kanamycin
D / 6	pOPS0532	Kanamycin
D / 7	pOPS0533	Kanamycin
D / 8	pOPS0534	Kanamycin
D / 9	pOPS0535	Kanamycin
D / 10	pOPS0536	Kanamycin
D / 11	pOPS0537	Kanamycin
D / 12	pOPS0538	Kanamycin
E / 1	pOPS0539	Kanamycin
E / 2	pOPS0540	Kanamycin
E / 3	pOPS0541	Kanamycin
E / 4	pOPS0542	Kanamycin
E / 5	pOPS0543	Kanamycin
E / 6	pOPS0544	Kanamycin
E / 7	pOPS0545	Kanamycin
E / 8	pOPS0546	Kanamycin
E / 9	pOPS0547	Kanamycin
E / 10	pOPS0548	Kanamycin
E / 11	pOPS0549	Kanamycin
E / 12	pOPS0550	Kanamycin
F / 1	pOPS0551	Kanamycin
F / 2	pOPS0552	Kanamycin
F / 3	pOPS0553	Kanamycin
F / 4	pOPS0554	Kanamycin
F / 5	pOPS0555	Kanamycin
F / 6	pOPS0556	Kanamycin
F / 7	pOPS0557	Kanamycin
F / 8	pOPS0558	Kanamycin
F / 9	pOPS0559	Kanamycin
F / 10	pOPS0560	Kanamycin
F / 11	pOPS0561	Kanamycin
F / 12	pOPS0562	Kanamycin
G / 1	pOPS0563	Kanamycin
G / 2	pOPS0564	Kanamycin
G / 3	pOPS0565	Kanamycin
G / 4	pOPS0566	Kanamycin
G / 5	pOPS0567	Kanamycin
G / 6	pOPS0568	Kanamycin
G / 7	pOPS0569	Kanamycin
G / 8	pOPS0570	Kanamycin
G / 9	pOPS0571	Kanamycin
G / 10	pOPS0572	Kanamycin
G / 11	pOPS0573	Kanamycin
G / 12	pOPS0574	Kanamycin
H / 1	pOPS0575	Kanamycin
H / 2	pOPS0576	Kanamycin
H / 3	pOPS0577	Kanamycin
H / 4	pOPS0578	Kanamycin
H / 5	pOPS0579	Kanamycin
H / 6	pOPS0580	Kanamycin
H / 7	pOPS0581	Kanamycin
H / 8	pOPS0582	Kanamycin
H / 10	pOPS0584	Kanamycin
H / 11	pOPS0585	Kanamycin
H / 12	pOPS0586	Kanamycin

Data calculated @ 2024-04-19