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Cultivarium POSSUM Toolkit
(Kit # 1000000234 )

Depositing Lab:   Nili Ostrov, Charlie Gilbert, Cultivarium Tools

The POSSUM Toolkit (Plasmid Origins and Selectable markerS for Undomesticated Microbes) is a Golden Gate genetic toolkit developed to help researchers identify functional plasmids and establish genetic tractability in non-model microbes. The kit includes plasmid origins of replication and selection markers for a variety of bacterial and fungal species and other genetic parts for engineering non-model microbes.

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

Add to Cart
$ 640 USD + shipping

Available to academics and nonprofits only.

Original Publication

A scalable framework for high-throughput identification of functional origins of replication in non-model bacteria Gilbert C, Brumwell SL, Crits-Christoph A, Kang SC, Martin-Moldes Z, Alsharif W, Esmurria A, Nguyen M-A, Lee HH, Ostrov N. bioRxiv 2023.05.19.541510. doi: 10.1101/2023.05.19.541510. Article (Link opens in a new window).

Description

Establishing plasmid transformation is a fundamental step in the genetic manipulation of microorganisms, underpinning heterologous protein expression, genome engineering, gene editing, and other applications. Screening for functional origin of replication (ORI) sequences and selection markers is essential to this process, but there is a lack of standardized, scalable resources to achieve this in non-model microbes. To make this process simpler, cheaper, faster, and more accessible to researchers, we have developed the Plasmid Origins and Selectable markerS for Undomesticated Microbes (POSSUM) Toolkit. The kit provides 111 Level 0 plasmids, including ORIs and selection markers for a range of bacterial and fungal species, and 22 destination vectors, as well as all the components required to assemble them together for multiplexed plasmid screening in non-model microbes. These parts are compatible with dozens of other Golden Gate toolkits.

Schematics of kit plasmids and overview. "Level 0 part collection" includes: oriT, promoter-RBS, inducible promoter, NGS barcode, ORI, promoter, RBS, CDS, and terminator parts. These parts are combined via BsaI golden gate assembly into "Level 1 assemblies": Odd1, Odd2, Odd3, and Odd4. These assemblies are further combined via SapI golden gate assembly into "Level 2 assemblies": Even1, Even2, Even3, Even4. Overview text is as described in figure legend and Description sections.
  • Figure 1: Overview of the POSSUM Toolkit (left). Illustration of the Loop Assembly method employed by the POSSUM Toolkit (right). Alternating rounds of BsaI and SapI Golden Gate assemblies combine up to four parts at a time into destination vectors. Assembly levels alternate between Odd and Even destination vectors. oriT: conjugation origin of transfer; RBS: ribosome binding site; CDS: coding sequence; NGS: next-generation sequencing. Possum image generated using MidJourney.

Kit Documentation

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 Cultivarium POSSUM Toolkit was a gift from Nili Ostrov, Charlie Gilbert, and Cultivarium Tools (Addgene kit #1000000234).”

For your Reference section:

A scalable framework for high-throughput identification of functional origins of replication in non-model bacteria Gilbert C, Brumwell SL, Crits-Christoph A, Kang SC, Martin-Moldes Z, Alsharif W, Esmurria A, Nguyen M-A, Lee HH, Ostrov N. bioRxiv 2023.05.19.541510. doi: 10.1101/2023.05.19.541510. Article (Link opens in a new window).

Cultivarium POSSUM Toolkit - #1000000234

Resistance Color Key

Each circle corresponds to a specific antibiotic resistance in the kit plate map wells.

Inventory

Searchable and sortable table of all plasmids in kit. The Well column links to its kit plate map on the right, as well as lists the plasmid well location in its plate. The Plasmid column links to a plasmid's individual web page.

Kit Plate Map

96-well plate map for plasmid layout. Hovering over a well reveals the plasmid name, while clicking on a well opens the plasmid page.

Resistance Color Key

Chloramphenicol
Chloramphenicol and Gentamicin
Kanamycin
Ampicillin
Chloramphenicol and Kanamycin
Chloramphenicol and Tetracycline
Chloramphenicol and Bleocin (Zeocin)
Chloramphenicol and Erythromycin
Spectinomycin
Chloramphenicol and Ampicillin

Inventory

Well Plasmid Resistance
Plate 1 / A / 1 pGL0_1 [RK2 ORI]
Chloramphenicol
Plate 1 / A / 2 pGL0_2 [pBBR1 ORI]
Chloramphenicol
Plate 1 / A / 3 pGL0_3 [RSF1010 ORI]
Chloramphenicol
Plate 1 / A / 4 pGL0_4 [pIJ101 ORI]
Chloramphenicol
Plate 1 / A / 5 pGL0_5 [pAL5000 ORI]
Chloramphenicol
Plate 1 / A / 6 pGL0_7 [pSa ORI]
Chloramphenicol
Plate 1 / A / 7 pGL0_9 [pUB110 ORI]
Chloramphenicol
Plate 1 / A / 8 pGL0_11 [pWKS1 ORI]
Chloramphenicol
Plate 1 / A / 9 pGL0_12 [pAMβ1 ORI]
Chloramphenicol
Plate 1 / A / 10 pGL0_13 [pIP501 ORI]
Chloramphenicol
Plate 1 / A / 11 pGL0_15 [CEN6/ARSH4 ORI]
Chloramphenicol
Plate 1 / A / 12 pGL0_16 [2μ ORI]
Chloramphenicol
Plate 1 / B / 1 pGLEL_6 [EF]
Chloramphenicol
Plate 1 / B / 2 pGL0_18 [AMA1 long ORI]
Chloramphenicol
Plate 1 / B / 3 pGL0_19 [panARS ORI]
Chloramphenicol
Plate 1 / B / 4 pGL0_20 [panARS-opt ORI]
Chloramphenicol
Plate 1 / B / 5 pGL0_21 [UARS1 ORI]
Chloramphenicol
Plate 1 / B / 6 pGL0_22 [pSG5 ORI]
Chloramphenicol
Plate 1 / B / 7 pGL0_23 [pSC101ts ORI]
Chloramphenicol
Plate 1 / B / 8 pGL0_24 [pBBR1-UP ORI]
Chloramphenicol
Plate 1 / B / 9 pGL0_25 [P3 promoter]
Chloramphenicol
Plate 1 / B / 10 pGL0_26 [bla pBR322]
Chloramphenicol
Plate 1 / B / 11 pGL0_27 [Tfd gene VIII]
Chloramphenicol
Plate 1 / B / 12 pGL0_28 [Paph(3')-I EZ-Tn5]
Chloramphenicol
Plate 1 / C / 1 pGL0_29 [aph(3')-I EZ-Tn5]
Chloramphenicol
Plate 1 / C / 2 pGL0_30 [PaphA-3 ]
Chloramphenicol
Plate 1 / C / 3 pGL0_31 [aphA-3]
Chloramphenicol
Plate 1 / C / 4 pGL0_32 [Pcat]
Chloramphenicol
Plate 1 / C / 5 pGL0_33 [cat]
Chloramphenicol
Plate 1 / C / 6 pGL0_34 [PaadA]
Chloramphenicol
Plate 1 / C / 7 pGL0_35 [aadA]
Chloramphenicol
Plate 1 / C / 8 pGL0_36 [PtetA pSC101]
Chloramphenicol
Plate 1 / C / 9 pGL0_37 [tetA pSC101]
Chloramphenicol
Plate 1 / C / 10 pGL0_38 [Paac(3)-Ia]
Chloramphenicol
Plate 1 / C / 11 pGL0_39 [aac(3)-Ia]
Chloramphenicol
Plate 1 / C / 12 pGL0_40 [dfrA1]
Chloramphenicol
Plate 1 / D / 1 pGL0_41 [Paac3-IV]
Chloramphenicol
Plate 1 / D / 2 pGL0_42 [aac3-IV]
Chloramphenicol
Plate 1 / D / 3 pGL0_43 [tsr]
Chloramphenicol
Plate 1 / D / 4 pGL0_44 [hph]
Chloramphenicol
Plate 1 / D / 5 pGL0_45 [nat1]
Chloramphenicol
Plate 1 / D / 6 pGL0_46 [nptII]
Chloramphenicol
Plate 1 / D / 7 pGL0_47 [bar]
Chloramphenicol
Plate 1 / D / 8 pGL0_48 [ble]
Chloramphenicol
Plate 1 / D / 9 pGL0_49 [tk101]
Chloramphenicol
Plate 1 / D / 10 pGL0_50 [catE]
Chloramphenicol
Plate 1 / D / 11 pGL0_51 [PcatA]
Chloramphenicol
Plate 1 / D / 12 pGL0_52 [catA]
Chloramphenicol
Plate 1 / E / 1 pGL0_53 [TcatA]
Chloramphenicol
Plate 1 / E / 2 pGL0_54 [PermC]
Chloramphenicol
Plate 1 / E / 3 pGL0_55 [ermC]
Chloramphenicol
Plate 1 / E / 4 pGL0_56 [TermC]
Chloramphenicol
Plate 1 / E / 5 pGL0_57 [PtetL]
Chloramphenicol
Plate 1 / E / 6 pGL0_58 [tetL]
Chloramphenicol
Plate 1 / E / 7 pGL0_59 [TtetL]
Chloramphenicol
Plate 1 / E / 8 pGL0_60 [sfGFP]
Chloramphenicol
Plate 1 / E / 9 pGL0_62 [sfYFP]
Chloramphenicol
Plate 1 / E / 10 pGL0_64 [mScarlet]
Chloramphenicol
Plate 1 / E / 11 pGL0_65 [StayGold]
Chloramphenicol
Plate 1 / E / 12 pGL0_76 [ORI_BC_1]
Chloramphenicol
Plate 1 / F / 1 pGL0_77 [ORI_BC_2]
Chloramphenicol
Plate 1 / F / 2 pGL0_78 [ORI_BC_3]
Chloramphenicol
Plate 1 / F / 3 pGL0_79 [ORI_BC_4]
Chloramphenicol
Plate 1 / F / 4 pGL0_80 [ORI_BC_5]
Chloramphenicol
Plate 1 / F / 5 pGL0_81 [ORI_BC_6]
Chloramphenicol
Plate 1 / F / 6 pGL0_82 [ORI_BC_7]
Chloramphenicol
Plate 1 / F / 7 pGL0_83 [ORI_BC_8]
Chloramphenicol
Plate 1 / F / 8 pGL0_84 [ORI_BC_9]
Chloramphenicol
Plate 1 / F / 9 pGL0_85 [ORI_BC_10]
Chloramphenicol
Plate 1 / F / 10 pGL0_86 [ORI_BC_11]
Chloramphenicol
Plate 1 / F / 11 pGL0_87 [ORI_BC_12]
Chloramphenicol
Plate 1 / F / 12 pGL0_88 [ORI_BC_13]
Chloramphenicol
Plate 1 / G / 1 pGL0_89 [ORI_BC_14]
Chloramphenicol
Plate 1 / G / 2 pGL0_90 [ORI_BC_15]
Chloramphenicol
Plate 1 / G / 3 pGL0_91 [ORI_BC_16]
Chloramphenicol
Plate 1 / G / 4 pGL0_92 [ORI_BC_17]
Chloramphenicol
Plate 1 / G / 5 pGL0_93 [ORI_BC_18]
Chloramphenicol
Plate 1 / G / 6 pGL0_94 [ORI_BC_19]
Chloramphenicol
Plate 1 / G / 7 pGL0_95 [ORI_BC_20]
Chloramphenicol
Plate 1 / G / 8 pGL0_96 [J23100]
Chloramphenicol
Plate 1 / G / 9 pGL0_97 [J23108]
Chloramphenicol
Plate 1 / G / 10 pGL0_98 [J23114]
Chloramphenicol
Plate 1 / G / 11 pGL0_99 [bsRBS]
Chloramphenicol
Plate 1 / G / 12 pGL0_100 [Pbs]
Chloramphenicol
Plate 1 / H / 1 pGL0_101 [PnptII]
Chloramphenicol
Plate 1 / H / 2 pGL0_102 [pAgTEF1]
Chloramphenicol
Plate 1 / H / 3 pGL0_103 [tAgTEF1]
Chloramphenicol
Plate 1 / H / 4 pGL0_104 [pAnTrpC]
Chloramphenicol
Plate 1 / H / 5 pGL0_105 [tAnTrpC]
Chloramphenicol
Plate 1 / H / 6 pGL0_106 [TAmarelleT1]
Chloramphenicol
Plate 1 / H / 7 pGL0_107 [TAmarelleT7]
Chloramphenicol
Plate 1 / H / 8 pGL0_108 [TAmarelleT9]
Chloramphenicol
Plate 1 / H / 9 pGL0_109 [TAmarelleT12]
Chloramphenicol
Plate 1 / H / 10 pGL0_110 [pNG2]
Chloramphenicol
Plate 1 / H / 11 pGL0_112 [oriT]
Chloramphenicol
Plate 1 / H / 12 pGL0_113 [Dummy ORI]
Chloramphenicol
Plate 2 / A / 1 pGL0_114 [p15A]
Chloramphenicol
Plate 2 / A / 2 pGL0_115 [pBC1 ORI]
Chloramphenicol
Plate 2 / A / 3 pGL0_121 [ORI_BC_21]
Chloramphenicol
Plate 2 / A / 4 pGL0_122 [ORI_BC_22]
Chloramphenicol
Plate 2 / A / 5 pGL0_123 [ORI_BC_23]
Chloramphenicol
Plate 2 / A / 6 pGL0_124 [ORI_BC_24]
Chloramphenicol
Plate 2 / A / 7 pGL0_125 [ORI_BC_25]
Chloramphenicol
Plate 2 / A / 8 pGL0_126 [ORI_BC_26]
Chloramphenicol
Plate 2 / A / 9 pGL0_127 [ORI_BC_27]
Chloramphenicol
Plate 2 / A / 10 pGL0_128 [ORI_BC_28]
Chloramphenicol
Plate 2 / A / 11 pGL0_129 [ORI_BC_29]
Chloramphenicol
Plate 2 / A / 12 pGL0_130 [ORI_BC_30]
Chloramphenicol
Plate 2 / B / 1 pGL0_142 [EilR-PjexD]
Chloramphenicol
Plate 2 / B / 2 pGL0_143 [EilR-PjexH1]
Chloramphenicol
Plate 2 / B / 3 pGL0_144 [EilR-PjexH2]
Chloramphenicol
Plate 2 / B / 4 pGLEL_4 [BF]
Chloramphenicol
Plate 2 / B / 5 pGLEL_5 [CF]
Chloramphenicol
Plate 2 / B / 6 pGLDV_17 [380G]
Chloramphenicol
Plate 2 / B / 7 pGLDV_13 [38e1G]
Chloramphenicol
Plate 2 / B / 8 pGLDV_14 [38e2G]
Chloramphenicol
Plate 2 / B / 9 pGLDV_15 [38e3G]
Chloramphenicol
Plate 2 / B / 10 pGLDV_16 [38e4G]
Chloramphenicol
Plate 2 / B / 11 pGLDV_20 [31e1G]
Chloramphenicol
Plate 2 / B / 12 pGL2_141
Chloramphenicol
Plate 2 / C / 1 pGL2_144
Chloramphenicol
Plate 2 / C / 2 pGL1_7
Kanamycin
Plate 2 / C / 3 pGL1_8
Kanamycin
Plate 2 / C / 4 pGL1_9
Kanamycin
Plate 2 / C / 5 pGL1_35
Kanamycin
Plate 2 / C / 6 pGL1_36
Kanamycin
Plate 2 / C / 7 pGL1_37
Kanamycin
Plate 2 / C / 8 pGL1_38
Kanamycin
Plate 2 / C / 9 pGL1_39
Kanamycin
Plate 2 / C / 10 pGL1_40
Kanamycin
Plate 2 / C / 11 pGL1_41
Kanamycin
Plate 2 / C / 12 pGL1_42
Kanamycin
Plate 2 / D / 1 pGL1_43
Kanamycin
Plate 2 / D / 2 pGL1_44
Kanamycin
Plate 2 / D / 3 pGL1_45
Kanamycin
Plate 2 / D / 4 pGL1_46
Kanamycin
Plate 2 / D / 5 pGL1_47
Kanamycin
Plate 2 / D / 6 pGL1_48
Kanamycin
Plate 2 / D / 7 pGL1_59
Kanamycin
Plate 2 / D / 8 pGLEL_1 [βω]
Kanamycin
Plate 2 / D / 9 pGLEL_2 [γω]
Kanamycin
Plate 2 / D / 10 pGLEL_3 [εω]
Kanamycin
Plate 2 / D / 11 pGLDV_21 [28o1*G]
Kanamycin
Plate 2 / D / 12 pGLDO_1 [α-sfYFP-β]
Kanamycin
Plate 2 / E / 1 pGL1_1
Kanamycin
Plate 2 / E / 2 pGL1_2
Kanamycin
Plate 2 / E / 3 pGL1_3
Kanamycin
Plate 2 / E / 4 pGLDV_5 [28o1G]
Kanamycin
Plate 2 / E / 5 pGLDV_6 [28o2G]
Kanamycin
Plate 2 / E / 6 pGLDV_7 [28o3G]
Kanamycin
Plate 2 / E / 7 pGLDV_8 [28o4G]
Kanamycin
Plate 2 / E / 8 pGLDV_42 [21o1G]
Kanamycin
Plate 2 / E / 9 pGL1_4
Kanamycin
Plate 2 / E / 10 pGL1_5
Kanamycin
Plate 2 / E / 11 pGL1_6
Kanamycin
Plate 2 / E / 12 pGLDV_10 [18e2G]
Ampicillin
Plate 2 / F / 1 pGLDV_11 [18e3G]
Ampicillin
Plate 2 / F / 2 pGLDV_12 [18e4G]
Ampicillin
Plate 2 / F / 3 pGLDV_19 [11e1G]
Ampicillin
Plate 2 / F / 4 pGLDV_1 [18o1G]
Ampicillin
Plate 2 / F / 5 pGLDV_2 [18o2G]
Ampicillin
Plate 2 / F / 6 pGLDV_3 [18o3G]
Ampicillin
Plate 2 / F / 7 pGLDV_4 [18o4G]
Ampicillin
Plate 2 / F / 8 pGLDV_41 [11o1G]
Ampicillin
Plate 2 / F / 9 pGLDV_9 [18e1G]
Ampicillin
Plate 2 / F / 10 pGL2_136
Chloramphenicol and Kanamycin
Plate 2 / F / 11 pGL2_137
Chloramphenicol and Tetracycline
Plate 2 / F / 12 pGL2_142
Chloramphenicol and Bleocin (Zeocin)
Plate 2 / G / 1 pGL2_145
Chloramphenicol and Erythromycin
Plate 2 / G / 2 pGL0_17 [AMA1 short ORI]
Spectinomycin
Plate 2 / G / 3 pGL2_135
Chloramphenicol and Ampicillin
Plate 2 / G / 4 pGL2_138
Chloramphenicol and Gentamicin
Plate 2 / G / 5 pGL2_175
Chloramphenicol and Gentamicin
Plate 2 / G / 6 pGL2_176
Chloramphenicol and Gentamicin
Plate 2 / G / 7 pGL2_177
Chloramphenicol and Gentamicin
Plate 2 / G / 8 pGL2_161
Chloramphenicol and Gentamicin
Plate 2 / G / 9 pGL2_162
Chloramphenicol and Gentamicin
Plate 2 / G / 10 pGL2_163
Chloramphenicol and Gentamicin
Plate 2 / G / 11 pGL2_164
Chloramphenicol and Gentamicin
Plate 2 / G / 12 pGL2_165
Chloramphenicol and Gentamicin
Plate 2 / H / 1 pGL2_166
Chloramphenicol and Gentamicin
Plate 2 / H / 2 pGL2_167
Chloramphenicol and Gentamicin
Plate 2 / H / 3 pGL2_168
Chloramphenicol and Gentamicin
Plate 2 / H / 4 pGL2_169
Chloramphenicol and Gentamicin
Plate 2 / H / 5 pGL2_170
Chloramphenicol and Gentamicin
Plate 2 / H / 6 pGL2_171
Chloramphenicol and Gentamicin
Plate 2 / H / 7 pGL2_172
Chloramphenicol and Gentamicin
Plate 2 / H / 8 pGL2_173
Chloramphenicol and Gentamicin
Plate 2 / H / 9 pGL2_174
Chloramphenicol and Gentamicin
Data calculated @ 2024-04-15

Kit Plate Map - #1000000234

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