Compressed genome set
Pricing and shipping information
The FGSC holds the full pLorist6Xh library used in the Neurospora genome program at the Broad Institute and can distribute individual clones or groups of clones. The full library is not available without special arrangement.
Pricing and shipping information
Orbach/Sachs cosmid library of N. crassa DNA sequences
(pMOcosX).
The vector pMOcosX has dominant selectable markers for fungi (hygromycin resistance)
and E. coli (ampicillin resistance). To prepare the library, pMOcosX was digested with
XbaI and the ends treated with phosphatase. The vector was then digested with XhoI
and the ends partially filled in with dCTP and dTTP using klenow. 74-OR23-1VA DNA
was partially digested with MboI and the ends partially filled in with dATP and dGTP
using klenow. The vector and genomic DNAs were ligated. The ligated DNA was
divided into two parts. One part was packaged using Stratagene gold II extracts and the
other part was packaged using Stratagene XL II extracts. The packaged material was
plated on host strain DH5aMCR. 2400 colonies were picked from each packaged DNA
set to 25 microtiter plates so that the entire library consists of 50 microtiter plates.
A map of pMOcosX
A list of identified cosmids in the library. Walk Data
Pricing and shipping information
Return to the TOP of this page
This library was constructed using the cosmid pcosAX as a vector and partially digested chromosomal DNA extracted from a wild type strain (ORS-a, FGSC 2490) as inserts. Cosmid pcosAX (designed, constructed, and provided by Dr. John Hamer, Purdue University) is a pWEl 5 derivative that contains the argB gene from Aspergillus nidulans substituting both the origin of replication (SV40ORI) and the Neomycin resistance gene (Neor). In addition, pcosAX also contains an extra Xhol restriction site inserted in between the T3 and T7 RNA polymerase phage promoters. To construct the library, pcosAX was digested with Xhol, the ends of the vector were then partially filled-in with dCTP and dTTP using DNA polymerase I - Klenow fragment, and dephosphorylated with alkaline phosphatase. 2490 DNA was partially digested with Sau3A and the ends were partially filled-in with dATP and dGTP using DNA polymerase I - Klenow fragment. Vector and genomic DNAs were ligated and packaged using Gigapack XL extracts (Stratagene). The packaged material was plated on host Escherichia coli XLl Blue MR (Stratagene). 24,000 colonies, corresponding to the primary library were pooled together and stored at -70 C using standard procedures. The frozen pool contains approximately 3 X 109cfu/ml.
Pricing and shipping information
All clones are end-sequenced and as such can be identified on the Neurospora genome.
This library will be described in the Fungal Genetics Newsletter.
For more information about this library, please contact the FGSC.
cDNA libraries from the Functional Genomics Program
13 different libraries are available. They are described
here
They are sent as an aliquot of the phage suspension.
Pricing and shipping information
N. crassa cDNA libraries constructed by Matthew SachsTwo cDNA libraries, which represent mRNA from the Neurospora crassa wild type strain, 74-OR23-1VA, were constructed in Lambda Zap version I+ (Short, Fernandez, Sorge and Huse. 1988 Nucl. Acids Res. 16:7583-7600) as described (Orbach, Sachs and Yanofsky. 1990 J. Biol.Chem. 265:10981-10987). cDNA libraries were prepared from macroconidia incubated in 1X Vogel's minimal medium with 2% sucrose for 0.5, 1, and 2 hours (combined to obtain the germinating library) and 6 hours (mycelial library). The mycelial library is estimated to contain 3.25 x 10^5 independent clones (75% recombinant). The libraries are supplied as lambda phage in SM containing 7% DMSO. A working aliquot of the library may be stored at 4 C. The remainder should be quick frozen in liquid nitrogen and stored below -70 C.
The germinating conidia library is no longer available.
Pricing and shipping information
Return to the TOP of this page
Nitrogen induced cDNA libraries constructed by R.H. Garrett
One library (NO3- induced) was made from poly(A)+ RNA isolated from Neurospora
crassa mycelia grown on Vogel's minimal made with 3% sucrose and
20 mM glutamine, then induced for nitrate assimilation by transfer to a
medium containing 20 mM nitrate as the sole N-source. The other library
(glutamine-grown) was made from poly(A)+ RNA isolated from mycelia grown as
above, then transferred to a medium containing 20 mM glutamine as the sole
N-source. cDNA in each library was prepared from poly(A)+ RNA using the
Pharmacia cDNA kit, ligated on EcoRI adapters, and cloned into lambda ZAP
version II+ Both libraries are >98% recombinant phage as judged by
blue/white selection and by individual plaque isolation.
Pricing and shipping information
An on-line order form
Return to the TOP of this page
2. Mycelial library: M-1, constructed with mRNA from conidia of 74A grown in 1X Vogel's and 2% sucrose, shaking at 25 C for 24 hours
3. Mycelial library (Westergaard's): constructed with mRNA from conidia of 74A grown in 1X Westergaard's and 1% sucrose, floating at 25 C for 1.5 days
4. Perithecial library: P-1, constructed with mRNA from 5 day-old perithecia [the fluffy a strain (FGSC #4347) was grown on crossing plates covered with sterile Miracloth circles and fertilized with a heavy conidial suspension of the opposite mating type (74A)---> day 0]
The libraries were constructed using the Uni-ZAP XR vector system
(Stratagene). Detailed description of screening and manipulation of the
libraries can be found in a Stratagene's instruction manual. The
bacterial strain XL1-blueMRF' can be used to plate the libraries. To
excise the pBluescript containing the cDNA from the library, a helper phage
(ExAssist) and an E. coli(SOLR) are needed.
Described in:
Expressed Sequences from Conidial, Mycelial, and Sexual Stages of Neurospora
crassa. Mary Anne Nelson, et al.Fungal Genetics and Biology Volume 21, Issue
3, June 1997, Pages 348-363
Please see the information about restrictions on
distribution of zap based libraries.
Return to the TOP of this page
+ Lambda ZAP versions I, II and Uni-ZAP XR are proprietary vectors owned by
Stratagene Cloning Systems.
Stratagene has kindly allowed the FGSC to distribute these libraries for
research purposes only to non-profit institutions. These libraries may
not be used for commercial purposes nor may they be used to reconstitute
the lambda zap vectors. Request for and acceptance of these libraries
constitutes acceptance of the following terms: The Lambda ZAP vectors shall
not be used for the reproduction, amplification or modification of the
vector. Neither the Lambda ZAP vectors nor derivatives of them shall be
offered for resale. Neither the Lambda ZAP vectors nor derivatives of them
shall be distributed or transferred to third parties.
For more information, send us an E-mail
or
Return to the FGSC main page