Generic sample from Heterodera glycines
Heterodera glycines
The genomic DNA sequence comes from the nematode Heterodera glycines, the common name is the soybean cyst nematode. DNA sequence data collected from strain of the nematode, called TN20.i Nematode DNA was extracted from frozen second-stage juveniles of the soybean cyst nematode, species Heterodera glycines. A standard Proteinase K with phenol-chloroform extraction method was used (see details below). SCN eggs were harvested as described in Niblack (1993). Eggs were deposited on top of an egg hatchi ng apparatus consisting of 41 micron mesh sieve held taut between 2 stacked 50 ml polypropylene bea kers with there bottoms cut off. The bottom surface of the sieve cloth was carefully laid on top of  a shallow layer of sterile dH20 containing 1%streptomycin sulfate solution (Sigma) in a 60mm petri  dish bottom. As the second-stage juveniles (J2s) hatched, they crawled through the mesh to the liq uid in the petri dish. The clean J2s were collected from the petri dish after allowing the J2 to ac cumulate for 24 hours. The J2s were concentrated by centrifugation until a 250 ul packed J2 pellet was obtained, then the J2's were flash frozen in liquid nitrogen. The frozen J2 pellet was transferred to a tissue publverizer (Fisher Scientific) precooled in liqui d nitrogen and crushed.  The pulverized frozen nematode fragments were mixed with 2 ml of proteinas e K solution (0.1M Tris-HCl (pH 8.0), 1 mM EDTA, 0.15M NaCl, 1% sodium dodecyl sulfate, 1mM dithiot hreitol, 2 mg/ml proteinase K) and were incubated for one hour at 37C.  The DNA was phenol/chlorofo rm-extracted, then ethanol precipitated as described in (Sambrook et al. 1989). The DNA was resuspe nded in 1 ml of TE (10 mM Tris-HCl pH 8, EDTA pH 8) containing RNase A at 10 microgram per ml and i ncubated at 37 degrees C for one hour. The DNA was next diluted to 20ml with column buffer (750 mM  NaCl; 50mM MOPS , pH 7; 15% isopropanol, 0.15% Triton X-100) and then further purified on a Genomic -tip 500/G as described in the manufacturers protocol (Qiagen, Valencia, CA).  The eluted SCN genom ic DNA was concentrated by ethanol precipitation and resuspended in 200 ul of TE.
STRAIN
TN20