Walbot Lab at Stanford University

Collaborations

We maintain our annual summer project at Cal Poly SLO, now in its fifth year in 2009. This involves 3 or 4 undergraduates each summer in our field screen for new Mu-tagged mutants of key genes in anther development. See the Methods section for a handbook describing this summer research experience.

Dave Skibbe of Stanford (second from left) smiles for the camera, reflecting the excellent help he received from Sarah (far left), Eric, Cedric, and Kristen in dissecting tens of thousands of staged anthers from Mutator tassels. Multiple replicate samples of each stage from four pairs of sister active and inactive Mutator lines are being used in proteomics and transcriptome profiling to pinpoint gene expression changes during anther development and to discover proteins whose abundance or post-translational status is affected by Mutator activity.

Our 2005 Stanford field hat commemorates the drive to Cal Poly -- 3 hours on Highway 101 -- well worth every minute to work with our collaborators at SLO.

Anther Cell Fate Setting Projects:
Cande lab collaboration. The anther project is a joint effort with Zac Cande’s lab at UC-Berkeley. Most of our current projects derive from their initial cytological screen to identify mutants that affect the earliest stages of anther development, during cell proliferation and cell fate setting. The charts summarize our current 2009 activities.

Locus	Gene tagging and cloning progress.	Help from
am1	CLONED Mu TE tagging Cande lab Switch from mitosis to meiosis Cande lab
msca1	CLONED Mu TE tagging & deletions Turn off leaf development program Dave Skibbe	Pioneer and ms6064* Dave working on this
mac1	CLONED Arabidopsis/rice candidates queried against maize map position then PCR screening of reference and Mu tagged alleles Meiotic cell fate Rachel, Tim, Dave, Gillian, Inna	proof that the MuDR insertion site causes mac1 phenotype
ms8	Getting close Arabidopsis candidates queried against maize map position then PCR screening of reference and Mu tagging alleles Pace of cell division & expansion; specification of meiotic cells Dong Xue	mtm99-56 from CSHL
ms23	Cloning in progress 7 new Mu tagged alleles One extra tapetal cell division Dave Skibbe working on this	historic alleles from Coop Dave working on this
ms32	Cloning in progress using new Mu tagged plus historic allele One extra tapetal cell division. More tagging summer 2009. Dave Skibbe working on this	historic Coop allele plus ms6066*
ARF2	CLONED Candidate gene had a RescueMu sequenced insertion sites Meiotic cell fate and anther structure Dong Xue	Reverse genetics from PCR row & column screening of grids
ms25	Cloning in progress – 3 Mu tagged alleles Vacuolate tapetal cells; microspores die Srividya working on this
mei775	Cloning in progress “gene discovery” – 3 Mu tagging alleles Pre-meiotic callose defect and then meiotic failure Dong Xue working on this	Another screen summer 2009
ms00-66*	“gene discovery” Tagging populations to screen summer 2009 Very small anthers	1 “iffy” Mu mutant now plus original allele

Allelism testing in progress (note some promising cases turned out to be alleles of known genes) and some tagging populations already done.

“Locus”	Key phenotype	Allelism test to all lines	Mu Tagging	Notes
ms6066* DROPPED	“gene discovery” Collapse of meiocytes; extra tapetal divisions	Hawaii 08 Score SLO 08	Hawaii 08 Screen SLO 08	Allele of ms32
mtm99-56 DROPPED	“gene discovery” Collapse of meiocytes and tapetum after normal layers form	Hawaii 08 Score SLO 08	Hawaii 08 Screen SLO 08	Allele of ms8
ms6064* DROPPED	“gene discovery” No meiocytes or other cell types	Hawaii 08 Score SLO 08	Hawaii 08 Screen SLO 08	*msca1* allele
ms9	Irregular tapetum	09	2010	?mtm99-66
ms10	Tapetal cells blow up then meiocytes fail	09	2010
ms11	Tapetal failure	09	2010	Some similarity to ms8
ms13	Excess anticlinal and periclinal tapetal divisions	2010	2011
ms14	Small and then disappearing meiocytes	09	2010

2009 Mu tagging populations
“Locus”	Key phenotype	Allelism test to all lines	Mu Tagging	Notes – allelism tests
ms6015*	“gene discovery” Extra tapetal cell divisions	Hawaii 08 Score SLO 09	Stanford 09 Screen SLO 10
ms2499*	“gene discovery” Very small anthers	Hawaii 08 Score SLO 09	Stanford 09 Screen SLO 10
EMS-63089	“gene discovery” No tapetum	Hawaii 08 Score SLO 09	Stanford 09 Screen SLO 10
tcl1	“gene discovery” Tapetum continues anti & periclinal divisions		Stanford 09 Screen SLO 10	Not mac1 Be sure to test with ms*6015
ms10	Tapetal cells blow up then meiocytes fail	09	Stanford 09 Screen SLO 10
ms11	Tapetal failure	09	Stanford 09 Screen SLO 10	Some similarity to ms8
ms14	Small and then disappearing meiocytes	09	Stanford 09 Screen SLO 10

Small RNAs during Anther Development. We are keenly interested in the small RNA populations of developing anthers and collaborate with Lew Bowman (Univ. South Carolina) and Sundaresan (UC-Davis) and their laboratories to analyze these molecules after deep sequencing.

Proteomics. Working with Al Burlingame’s lab at UC-SF we are identifying proteins differentially expressed during anther development, whether through de novo synthesis or post-translational modifications. Currently this project is performed by postdocs Dong Xue Wang and Dave Skibbe. We also plan to conduct small proteome analyses on tassel proteins to look for potential signalling peptides.