Journal article
Fluorescent and bimolecular-fluorescent protein tagging of genes at their native loci in Neurospora crassa using specialized double-joint PCR plasmids
Fungal genetics and biology, Vol.48(9), pp.866-873
2011
DOI: 10.1016/j.fgb.2011.05.002
PMID: 21664475
Abstract
► Novel plasmids created for targeted genome integration in
Neurospora crassa. ► Designed for GFP, RFP, and split-YFP (BiFC) tagging. ► Customized construction involves simple DJ-PCR steps. ► Various meiotic silencing proteins have been successfully tagged with this technique. ► SMS-2 and QIP have been shown to interact in the perinuclear region.
The double-joint polymerase chain reaction (DJ-PCR) is a technique that can be used to construct vectors for targeted genome integration without laborious subcloning steps. Here we report the availability of plasmids that facilitate DJ-PCR-based construction of
Neurospora crassa tagging vectors. These plasmids allow the creation of green or red fluorescent protein (GFP or RFP) tagging vectors for protein localization studies, as well as split-yellow fluorescent protein (YFP) tagging vectors for bimolecular fluorescence complementation (BiFC) analyses. We have demonstrated the utility of each plasmid with the tagging of known meiotic silencing proteins. Microscopic analysis of the tagged strains indicates that SMS-2 and QIP form macromolecular complexes in the perinuclear region during meiosis.
Details
- Title: Subtitle
- Fluorescent and bimolecular-fluorescent protein tagging of genes at their native loci in Neurospora crassa using specialized double-joint PCR plasmids
- Creators
- Thomas M Hammond - Division of Biological Sciences, University of Missouri, Columbia, MO 65211, United StatesHua Xiao - Division of Biological Sciences, University of Missouri, Columbia, MO 65211, United StatesDavid G Rehard - Division of Biological Sciences, University of Missouri, Columbia, MO 65211, United StatesErin C Boone - Division of Biological Sciences, University of Missouri, Columbia, MO 65211, United StatesTony D Perdue - Department of Biology, University of North Carolina, Chapel Hill, NC 27599, United StatesPatricia J Pukkila - Department of Biology, University of North Carolina, Chapel Hill, NC 27599, United StatesPatrick K.T Shiu - Division of Biological Sciences, University of Missouri, Columbia, MO 65211, United States
- Resource Type
- Journal article
- Publication Details
- Fungal genetics and biology, Vol.48(9), pp.866-873
- Publisher
- Elsevier Inc
- DOI
- 10.1016/j.fgb.2011.05.002
- PMID
- 21664475
- ISSN
- 1087-1845
- eISSN
- 1096-0937
- Language
- English
- Date published
- 2011
- Academic Unit
- Biology
- Record Identifier
- 9984083275202771
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