The Karyotype Analysis Of Seven Strains From Four Species Of Entomogenous Fungi In Paecilomyces Bainier

The conditions of mycelia culturing, enzymolysis, protoplast preparation, electrophoresis and molecular karyotype analysis of seven strains from four species of entomogenous fungi in Paecilomyces Bainier were investigated, in order to obtain optimzed preparation of protoplasts suspension. In addition, the optimal electrophoresis condition which adapts to chromosomes DNA electrophoresis of entomogenous fungi was investigated with switching angle, voltage, electrophoresis time, pulse intervals. The analysis of chromosomes and molecular karyotype of four species was carried out by using the CHEF system( Bio-Rad CHEF Mapper, CHEF DRIII, USA )with Hansenula wingei YB-4662-VIA ( Bio-Rad, Richmond, USA ) and Schizosaccharomyces pombe 972h- ( Bio-Rad, Richmond, USA ) chromosomes DNA as molecular weight markers, and CHEF patterns obtained were analyzed by the AlphaEase Fc ( Alphalmager 2200 ) software. Some original results were concluded as following.1. The protoplasts suspension was obtained with higher concentration ( 1.0×107/ml ) under the following conditions: the mycelia cultured for 18h, digested by the mixed enzyme solution consisting of 1% lywallzyme, 1% cellulase and 1% snailase while shaking by 120r/min for 2.5h at 30℃. The results of electrophoresis showed the suspension concentration of protoplasts varied from 1.9×108 to 3.8×108/ml less than 9.6×108/ml, in order to count the number and molecular weight of chromosomes and genome.2. The karyotype and chromosomes of two strains of Paecilomyces tenuipes were studied. One from body of Cordyceps takaomontana has eight chromosomes from 0.43 to 5.78 mega base pairs (Mb), 0.43, 2.45, 2.55, 2.88, 3.28, 3.94, 4.70 and 5.78Mb, respetively, total genome sizes estimated to be 26.01Mb, while the other from the anamorph of Cordyceps takaomontana has 7 chromosomes as following: 0.44, 2.46, 2.72, 2.99, 3.50, 4.49 and 5.37Mb, total genome sizes estimated as 21.97Mb. So, the karyotype and chromosomes of the above strains of Paecilomyces tenuipes are obviously diferent.3. Paecilomyces cateniannulatus were first studied by CHEF electrophoresis, obtaining eight chromosomes from 0.43 to 5.78Mb, as following: 0.44, 1.51, 2.20, 4.38, 4.81, 5.16, 5.37 and 5.85Mb. The total genome sizes were estimated to be 28.52Mb by molecular weight markers.4. The karyotype and chromosomes of two strains of Paecilomyces cateniobliquus were studied. One has seven chromosomes from 1.58 to 7.52Mb, as following: 1.58, 4.20, 4.82, 5.40, 6.17, 6.74 and 7.52Mb, total genome sizes estimated to be 34.43Mb, while the other also has 7 chromosomes as following: 3.93, 4.95, 5.63, 6.64, 6.74, 7.14 and 7.52Mb, total genome sizes estimated as 34.32Mb. which shows the two strains have a little diference in karyotype.5. The CHEF electrophoresis conditions of Paecilomyces fumosoroseus were optimized and more chromosomes were obtained as following: eight chromosomes with molecular weight from 0.45 to 5.78Mb, including 0.45, 2.87, 3.46, 4.33, 5.07, 5.85, 6.86 and 7.62Mb. The total genome sizes were estimated to be 36.49Mb.6. The number and molecular weight of chromosomes and karyotype seven strains from four species of entomogenous fungi in Paecilomyces Bainier were firstly investigated. The results showed that the chromosomes and karyotype are not all homological among the same genuses or the same strains. It is a bit difficult to analyze the heredity distance or genetic relaitionship of different species of Paecilomyces because of less difference in the number of chromosome but more diversity in karyotype. In general, the chromosome and karyotype diversity among strains is obviously less than that among species of Paecilomyces.