Cloning of the DNA fragments reflecting the open reading frame I and II of the I-18 C gene of Chironomus tentans. III. Preparation of the inserts for ligation with the bluescript and the modification of the vector

• Autorzy: Borowicz B P
• Języki publikacji: EN

Abstrakty

EN

The technology applied for the cloning of the open reading frame (ORF) I and II reflecting DNA fragments of the I-18 C gene of Chironomus tentans in regard to the preparation of the inserts for ligation with the bluescript and the modification of this intermediate vector, is described. The ORF I reflecting DNA fragment prepared for ligation with the bluescript vector, had one end blunt (i.e. the 5’ end with the Nde I restriction site) and the second end was sticky (i.e. the 3’ end after the Bam HI digestion of this insert). Subsequently, the bluescript vector for this ligation had also one end blunt (i.e. the 5’ end with the Nde I restriction site) and the second end was sticky (i.e. the 3’ end after the Bam HI digestion). The ORF II reflecting DNA fragment prepared for the ligation had both ends blunt and so the vector.

PL

Opisano technologię zastosowaną do klonowania fragmentów DNA, odzwierciedlających translacyjną otwartą ramę odczytu I i II genu I-18 C w zakresie przygotowania wstawek do ligacji z plazmidem blueskrypt i niezbędnych modyfikacji wektora pośredniego. Fragment DNA odzwierciedlający otwartą ramę odczytu (ORF) I przygotowano do ligacji z wektorem blueskrypt w ten sposób, że fragment ten posiadał jeden koniec tępy (tj. koniec 5’ z miejscem restrykcyjnym dla Nde I) i drugi koniec lepki (tj. koniec 3’ po trawieniu tej wstawki Bam HI). Także wektor blueskrypt, użyty do ligacji ze wspomnianą sekwencją wstawki, miał jeden koniec tępy (tj. koniec 5’ z miejscem restrykcyjnym dla Nde I) i drugi koniec lepki (tj. koniec 3’ po trawieniu Bam HI). Fragment DNA odzwierciedlający ORF II, przygotowany do ligacji z wektorem blueskrypt, miał oba końce tępe i stosownie do tego oba końce wektora były też tępe.

Słowa kluczowe

EN

cloning; pET-3a vector expression; I-18 C gene; T7 RNA polymerase; Chironomus tentans; promotor system; polymerase chain reaction; bluescript vector; DNA fragment

• Wydawca: -
• Czasopismo: Journal of Plant Protection Research
• Rocznik: 1999
• Tom: 39
• Numer: 1
• Opis fizyczny: p.50-53

Twórcy

autor

Borowicz B P

• Institute of Plant Protection, Miczurina 20, 60-318 Poznan, Poland

Bibliografia

• 1. Borowicz B.P. 1995. Isolation of the DNA fragment reflecting the Open Reading Frame I of the I-18 C gene of Chironomus tentans by the Polymerase Chain Reaction. I. The technology for the preparation of the samples (in English). Prace Nauk. Inst. Ochr. Roślin 36 (1/2): 69-76.
• 2. Borowicz B.P. 1995. Isolation of the DNA fragment reflecting the Open Reading Frame I of the I-18 C gene of Chironomus tentans by the Polymerase Chain Reaction. II. The technology required to obtain the DNA fragment (in English). Prace Nauk. Inst. Ochr. Roślin 36 (1/2): 77-82.
• 3. Borowicz B.P. 1995. Isolation of the DNA fragment reflecting the Open Reading Frame I of the I-18 C gene of Chironomus tentans by the Polymerase Chain Reaction. III. The effect of the applied technologies (in English). Prace Nauk. Inst. Ochr. Roślin 36 (1/2) : 83-93.
• 4. Borowicz B.P. 1996. Isolation of the DNA fragment reflecting the Open Reading Frame II of the I-18 C gene of Chironomus tentans by the Polymerase Chain Reaction. I. The technology for the preparation of the primers. J. Plant Protection Res., 37 (1/2): 138-144.
• 5. Borowicz B.P. 1996. Isolation of the DNA fragment reflecting the Open Reading Frame II of the I-18 C gene of Chironomus tentans by the Polymerase Chain Reaction. II. The technology for the preparation of the template. J. Plant. Protection Res., 37 (1/2): 145-148.
• 6. Borowicz B .P. 1996. Isolation of the DNA fragment reflecting the Open Reading Frame II of the I-18 C gene of Chironomus tentans by the Polymerase Chain Reaction. III. The DNA amplification technology. J. Plant Protection Res., 37 (1/2): 149-155.
• 7. Borowicz B.P. 1996. lsolation of the DNA fragment reflecting the Open Reading Frame II of the I-18 C gene of Chironomus tentans by the Polymerase Chain Reaction. IV. The product of the applied technologies. J. Plant Protection Res., 37 (1/2): 156-164.
• 8. Borowicz B.P. 1996. Isolation of the DNA fragment reflecting the Open Reading Frame II of the I-18 C gene of Chironomus tentans by the Polymerase Chain Rcaction. V. Different mechanisms of regulation regarding the Open Reading Frames. J. Plant Protection Res., 37 (1/2): 165-171.
• 9. Del Sal G.M., Manfioletti G., Schneider C. 1988. A one-tube plasmid DNA mini-preparation suitable for sequencing. Nucl. Acids Res. 16: 9878 pp.
• 10. Heery D.M., Gannon F., Powell R. 1990. A simple method for subcloning DNA fragments from gel slices. Trends in Genet., 6: 173 pp.
• 11. Lezzi M., Lutz B., Dorsch-Haesler K. 1989 b. The ecdysterone-controlled chromosome region I-18 C in C. tentans: from cytology to molecular biology. Acta Biol. Debr. Oecol. Hung., 2: 129-139.
• 12. Sambrook J., Fritsch E.F., Maniatis T. 1989. Molecular Cloning. A laboratory manual. Second Edition. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, Books: 1-3.
• 13. Sambrook J., Fritsch E.F., Maniatis T. 1989. Plasmid Vectors (chapter 1) in Molecular Cloning. A laboratory manual. Second Edition, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, Book 1: 1.2-1.110.
• 14. Sambrook J., Fritsch E.F., Maniatis T. 1989. Expression of Cloned Genes in Escherichia coli (chapter 17) in Molecular Cloning. A laboratory manual. Second Edition. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, Book 3: 17.2-17.44.
• 15. Sambrook J., Fritsch E.F., Maniatis T. 1989. Commonly Used Techniques in Molecular Cloning (Appendix E) in Molecular Cloning. A laboratory manual. Second Edition. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, Book 3: E.5-E.7.
• 16. Studier F.W., Rosenberg A.H., Dunn J.J., Dubendorff J.W. 1990. Use of T7 RNA Polymerase to direct expression of cloned genes. Methods in Enzymology, Goeddel D.V. (ed.), 1985: 60-89.
• 17. Winnacker E.-L. 1987. From Genes to Clones. lntroduction to Gene Technology (in English). VCH Verlags-gesellschaft mbH, D-6940 Weinheim, Germany: 1-634.