Krisana Phucharoen¹, Wipa Chungjatupornchai² and Sakol Panyim^1,2*

¹Institute of Molecular Biology and Genetics, Mahidol University, Salaya Campus,
Nakornpathom, Thailand; ¹National Center for Genetic Engineering and Biotechnology (BIOTEC), Bangkok, Thailand.

(Received 29 December 1998 / Accepted 26 April 1999)

Abstract.
A total of 29 subspecies of B. thuringiensis strains previously used to raise 28 standard antisera, were differentiated using PCR fingerprinting with primers derived from the Repetitive Extragenic Palindrome (REP) sequence. Using REP specific primers (REP2-1 and REP3) in a defined PCR condition, a reproducible fingerprinting pattern in the size range of 300 2,000 bp was obtained. The bacterial strains can be distinguished by comparing fingerprint patterns. The dendrogram of the fingerprints revealed that the B. thuringiensis subspecies fell into four major clusters with a level of similarity of approximately 20%. The relatedness of the REP-PCR fingerprint patterns of most B. thuringiensis subspecies did not correlate with the bacterial classification based on flagella serotyping. The REP-PCR fingerprint method should prove to be a rapid and simple tool for numerical taxonomic analysis of B. thuringiensis strains.

Keywords: Bacillus thuringiensis, REP PCR, genomic Fingerprintin

N. N. Hasima^*,1. Azli and A. A. Rahmat

Institute of Biological Sciences, Faculty of Science, University of Malaya, 50603 Kuala Lumpur, Malaysia

(Received 19February 1999 / Accepted 19 April 1999)

Abstract.
DNA fingerprints of chickens were successfully obtained using a non radioactive technique. The probe used was the human minisatellite probe, 33.15 and digestions of the genomic DNA were with HaeIII Hinfl and AluI. Average DNA fingerprint band sharing and band frequency between individuals with respect to the different proberestriction enzyme combinations were used to estimate average band sharing between individuals and average band frequency.

Keywords: Chicken, minisatellite, non radioactive labeling

Chanikarn Boonchuoy¹, Boonyaruk Boonyawan², Sakol Panyim²,
and Burachai Sonthayanon^*1,2

¹Central Equipment Laboratory, Institute of Science and Technology for Research and Development, Mahidol University,
Salaya Campus, and ²Institute of Molecular Biology and Genetics, Mahidol University, Salaya Campus,
Phutthamonthon 4 Rd., Phutthamonthon District, Nakhon Pathom 73170, Thailand

(Received 17 December 1998 / Accepted 23 March 1999)

Abstract.
A sequence determination was performed on a 1.86. kb cDNA clone designated as PMM020. The clone was among a set of random cDNA clones isolated from an abdominal muscle cDNA library of Black Tiger Prawn which had been partially sequenced for expressed sequence tags (5' EST) markers. Earlier database query via a BLAST program indicated that a partial DNA sequence from this clone matched enolase sequences from other eukaryotic organisms. DNA sequencing was thus performed on subcloned DNA fragments from PMM020 and an 1861 bp of combined sequence was found. A tract of poly (A) was found at the 3' side beyond position 1861 of the sequence, which indicated that the 3' end of the transcript was intact. An open reading frame for 434 amino acids was found starting from the predicted translation initiation codon (ATG) at nucleotide position 24 and ending at a termination codon (TAA) at position 1327. The predicted protein molecular weight was 47 kDa. An amino acid motif specific to enolase, DDLTVTNPK was found at residue positions 320 328. Upon comparing with an enolase cDNA sequence from Xenopus laevis, the overall nucleotide sequence identity between the two sequences was 62.0% while the identity within the open reading frame was 69.4% and the calculated protein sequence identity was 72.5%. When compared to those of other eukaryotes, the calculated amino acid sequence identities of 77.6% (Drosophila melanogaster), 63.1 % (yeast, Saccharomyces cerevisiae), 72.7% (chicken, Gallus gallus), 71.3% (mouse, Mus musculus) were found. The high percent identity for both nucleotide and predicted protein sequences, together with the expected length of the polypeptide chain, identified the cloned sequence as phosphopyruvate hydratase (GenBank accession no. AFI 00985)

Keywords: Prawn, phosphopyruvate hydratase, enolase, shrimp

Prasun K. Mukherjee

Nuclear Agriculture and Biotechnology Division, Bhabha Atomic Research Centre, Mumbai 400 085, India.

(Received 20 February 1999 / Accepted 7 April 1999)

Abstract.
A rapid method for plant genomic DNA extraction is described here. This method, which is a simple modification of the existing methods, requires only one centrifugation at I 1 000 g for 10 min. High molecular weight genomic DNA from various plants suitable for restriction analysis and PCR amplification has been obtained.

Keywords: DNA, plant, fungi, RFLP, RAPD