DNA cloning

Các kỹ thuật chính dùng trong công nghệ DNA tái tổ hợp 1.1. Khái niệm 1.2. Các enzym dùng trong công nghệ DNA tái tổ hợp 1.3. Các vector nhân dòng dùng trong công nghệ DNA tái tổ hợp 1.4. Nhân dòng gen (gene cloning) 1.5. Chọn lọc, tạo dòng và sự biểu hiện của gen 2. Các kỹ thuật chính sử dụng trong phân tích DNA 2.1. Kỹ thuật chiết tách DNA và RNA 2.2. Kỹ thuật tạo ngân hàng cDNA 2.3. Phương pháp PCR 2.4. Kỹ thuật xác định trình tự DNA 2.5. Kỹ thuật RFLP (dựa vào lai DNA/DNA) 2.6....

46p hoatuoi0939310 26-02-2013 286 81   Download

Việc chọn lựa phương pháp ly trích tùy thuộc vào nhiều yếu tố: Việ chọn lự pháp trích tù thuộ và nhiề yế tố – Nguồn ADN: từ máu, niêm mạc, VK, TV, ĐV Nguồ từ máu, mạ TV, – Mục đích sử dụng của ADN được ly trích: PCR, cắt bằng Mục đí sử dụng củ được trích: PCR, bằ Enz. cắt giới hạn, Cloning... giớ hạ – Loại ADN: DNA trong nhân (genomic DNA), ty thể, lạp thể Loại thể lạ thể – Số lượng ADN cần thu được, số mẫu cần ly trích ượng cầ được, cầ...

30p trada85 22-01-2013 137 25   Download

Bài giảng "Công nghệ di truyền - Chương 3: Simple cloning" cung cấp cho người học các kiến thức: Vector pUC, vai trò của các gene, cắt vector và DNA, nối bằng ligase được, chuyển tất cả cấu trúc vào E.coli, Operon lacZ và IPTG, chọn dòng mang DNA tái tổ hợp,... Mời các bạn cùng tham khảo nội dung chi tiết.

55p doinhugiobay_10 12-01-2016 112 19   Download

Bài giảng "Simple cloning" cung cấp cho người học các kiến thức: Vector pUC, vai trò của các gene, cấu trúc của các gen, cắt vector và DNA, nối bằng ligase được, chuyển tất cả cấu trúc vào E.coli, chọn dòng mang DNA tái tổ hợp, vector nhân tạo đầu tiên, tế bào chủ, tính chất của tế bào chủ, nhân dòng sản phẩm PCR,... Mời các bạn cùng tham khảo.

55p doinhugiobay_12 15-01-2016 57 5   Download

Bài giảng Sinh học phân tử: Chương 7 trình bày về kỹ thuật tạo dòng. Chương này trình bày những nội dung chính sau: Các công cụ sử dụng trong kỹ thuật tạo dòng, Restriction Enzyme (RE), vai trò sinh học của RE, restriction endonuclease, tạo dòng phân tử (molecular cloning), DNA tái tổ hợp, công cụ tạo dòng,... Mời các bạn tham khảo.

34p hihihaha2 03-12-2016 95 12   Download

Bài giảng Sinh học phân tử: Chương 7 Kỹ thuật tạo dòng, cung cấp cho người học những kiến thức như: Các công cụ sử dụng trong kỹ thuật tạo dòng; Vai trò sinh học của RE; Tạo dòng phân tử (molecular cloning); Sự khác nhau của các vector tạo dòng; Plasmid là các vector tạo dòng;...Mời các bạn cùng tham khảo!

60p caphesuadathemmatong 25-11-2021 29 2   Download

An optogenetic upgrade for the tet - OFF system fused the inhibitor to the blue light-responsive B-LID degron and optimized the performance of this construct with regard to the number of Med25VBD repeats. This approach resulted in an optogenetic upgrade of the popular Tet-OFF (TetR-VP64, tetO7-PhCMVmin)  system  that  allows tunable, blue light-inducible gene expression in HEK-293T  cells.

5p thuyancn 01-06-2015 30 5   Download

Telomerase is a specialized reverse transcriptase responsible for synthesizing telomeric DNA at the ends of chromosomes. Six subunits composing the telomerase complex have been cloned: hTR (human telomerase RNA), TEP1 (telomerase-associated protein 1), hTERT (human telomerase reverse transcriptase), hsp90 (heat shock protein 90), p23, and dyskerin. In this study, we investigated the role of each the telomerase subunit on the activity of telomerase.

9p research12 01-06-2013 28 3   Download

We report the cloning and functional charaterization of the full-length cDNA and gene encoding a Toxoplasma gondii DNA repair enzyme designated TgDRE. The gene is composed of three exons separated by two introns of 780 and 630 bp, and encodes a protein with a predicted molecular mass of 49.6 kDa. The native TgDRE protein, with a molecular mass of 60 kDa, is only detected in the virulent tachyzoite stage of T. gondii. However, the transcript is present in both asexual parasite stages, virulent tachyzoite and avirulent encysted bradyzoite. ...

9p research12 01-06-2013 36 5   Download

Primary structural information including the complete nucleotide sequence of the ®rst insect aldehyde oxidase (AO) was obtained from the common house mosquitoCulex quinquefasciatus(Say) through cloning and sequencing of both genomic DNA and cDNA. The deduced amino-acid sequence encodes a 150-kDa protein of 1266 amino-acid residues, which is consistent with the expected monomeric subunit size of AO.

12p research12 01-06-2013 49 5   Download

We have cloned the gene coding for theBacillus subtilis glycine oxidase (GO), a new ¯avoprotein that oxidizes gly-cine and sarcosine to the corresponding a-keto acid, ammonia and hydrogen peroxide. By inserting the DNA encoding for GO into the multiple cloning site of the expressionvectorpT7.7weproducedarecombinantplasmid (pT7-GO).ThepT7-GOencodes a fullyactive fusionprotein with six additional residues at the N-terminus of GO (MARIRA).

8p research12 29-04-2013 38 3   Download

The primary purpose of the present study was to investi-gate whether DNA replication at meiotic prophase also requires replication factors, especially proliferating cell nuclear antigen (PCNA). We cloned PCNA cDNAs (CoPCNA) from a cDNA library made from basidia of the basidiomycete,Coprinus cinereus. Interestingly, although CoPCNAis a single-copy gene in the genome, two di€er-entPCNAcDNA species were isolated using degenerate primers and a meiotic cDNA library, and were designated asCoPCNA-aandCoPCNA-b. ...

11p research12 29-04-2013 45 4   Download

Thea3b1 integrin is an adhesion receptor for extracellular matrix proteins including isoforms of laminin, and the changes of its expression level in various cancer cells are thought to cause their malignant phenotypes. We have cloned an approximately 4 kb DNA fragment of the 5¢-flanking region of the murinea3 integrin gene and analyzed its promoter activity.

9p research12 23-04-2013 37 3   Download

A nitroalkane-oxidizing enzyme gene (naoA) was cloned from a genomic DNA library ofStreptomyces ansochromo-genes7100. The deduced protein (NaoA) of this gene con-tains 363 amino acids and has high similarity to several nitroalkane-oxidizing enzymes from various micro-organ-isms.ThenaoAgenewas subcloned intoanexpressionvector pET23b and overexpressed inEscherichia coliBL21(DE3). The protein was then purified, and its characteristics were studied. Experimental results showed that NaoA can con-vert 1-nitropropane, 2-nitropropane and nitroethane into the corresponding carbonyl compounds....

6p tumor12 22-04-2013 44 2   Download

Human cytochrome P450 17a-hydroxylase (CYP17) cata-lyses not only the 17a-hydroxlation of pregnenolone and progesterone and the C17,20-side chain cleavage (lyase) of 17a-hydroxypregnenolone, necessary for the biosynthesis of C21-glucocorticoids and C19-androgens, but also catalyses the 16a-hydroxylation of progesterone. In efforts to under-stand the complex enzymology of CYP17, structure/func-tion relationships have been reported previously after expressingrecombinant DNAs, encodingCYP17 from various species, in nonsteroidogenic mammalian or yeast cells. ...

9p tumor12 22-04-2013 45 2   Download

Wehave identifiedanovelmammaliangene, termednicolin1 gene (NICN1), that is present in human, dog and mouse, whereas it is absent from the available genome sequences of nonmammalian organisms. TheNICN1gene consists of six exons and spans about 6 kb of genomic DNA. It encodes a 213 amino acid protein that does not belong to any known protein family. Experiments using green fluorescent protein (GFP)-taggednicolin1 fusionproteins indicate that nicolin1 is a nuclear protein.

6p tumor12 22-04-2013 53 2   Download

We have isolated and studied the cloned sigma factors SASIG1-3 from mustard (Sinapis alba). In functional ana-lyses using both promoter and factor mutants,the three recombinant proteins all had similar basic properties but also revealed differences in promoter preference and requirements for single nucleotide positions. Directedmuta-genesis of SASIG1 identified critical residues within the conserved regions 2.4 and 4.2 necessary for binding of the )10 and)35 promoter elements,respectively.

13p tumor12 20-04-2013 33 2   Download

We have previously cloned, expressedand characterized two variants of the major allergen Lep d 2 from culturedLepi-doglyphus destructor mites. These variants, Lep d 2.0101 and Lep d 2.0201, differ at 13 amino acid positions. In this study we investigated Lep d 2 sequence diversity between wild and cultured mites. PCR, Southern blot and DNA sequence analysis revealed the presence of two different Lep d 2 genes, one with and one without an intron.

8p tumor12 20-04-2013 49 3   Download

The physiological role of a bifunctional enzyme, 3,4-dihydrocoumarin hydrolase (DCH), which is capable of both hydrolysis of ester bonds and organic acid-assisted bromination of organic compounds, was investigated. Purified DCH fromAcinetobacter calcoaceticusF46 cata-lysed dose- and time-dependent degradation of peracetic acid. The gene (dch) was cloned from the chromosomal DNA of the bacterium.

9p tumor12 20-04-2013 39 3   Download

A complementary DNA encoding a new bovine tryptase isoform (here named BLT) was cloned and sequenced from lung tissue. Analysis of sequence indicates the pres-ence of a 26-amino acid prepro-sequence and a 245 amino acid catalytic domain. It contains six different residues when compared with the previously characterized tryptase from bovine liver capsule (BLCT), with the most signifi-cant difference residing at the primary specificity S1 pocket.

11p tumor12 20-04-2013 41 5   Download