Sequence Analysis Of Shiga Toxin 1 And Shiga Toxin 2 Genes Of Escherichia Coli O157: H7 Isikates From Lahore
By: Saqib Hussain
| Mr. Tanveer Hussain.
Contributor(s): Prof. Dr.Masroor Elahi Babar.
Material type: 
BookPublisher: 2011Subject(s): Institute of Biochemistry & BiotechnologyDDC classification: 1375,T Dissertation note: Escherichia coli is normal inhabitant of all the animals and human beings. The Sorbitol
non fermenting E. coli strains were detected in milk, beef and fecal samples collected
from different areas of Lahore. White colored colonies of each positive sample on
Sorbitol MacConkey's Agar (SMA) contained gram negative, rods with round ends non
spore former Escherichia coli. Each of the isolates was sorbitol non fermenter, lactose
fermenter, indole positive, Methyl Red positive, Voges Prauskaur negative and citrate
negative. Each of the isolate was further characterized using polymerase chain reaction
(PCR) for the presence of shiga toxin 1 and shiga toxin 2 genes. Bacterial DNA was
extracted easily by boiling method when the isolates were grown on Sorbitol
MacConkey's agar at 37°C for 12-24 hours. The DNA was recovered when the culture
was boiled for 5-10 minutes. The isolated DNA when amplified using Stxland Stx2
specific primers showed that 68.5 percent samples were positive for Stxl and 54.2
percent for Stx2. The stx 1 and stx2 PCR products were subjected to sequencing. The
resulted sequences when aligned with the reference sequence through Basic local
alignment tool it showed that the shiga toxin 1 and shiga toxin 2 gene sequences are
conserved and showed high similarity in their nucleotide structure. Despite of having
high similarity in their nucleotide structure some haplotypes were also obtained showing
single nucleotide polymorphism. Phylogenetic analysis made among local isolates and
also with reported sequences from all over the world by using bioinformatics software to
see the genetic similarities and difference between them. The data produced showed
some highly conserved sequences and SNPs as well that will be quite useful for further
applications in diagnostics and biotechnology applications in future.
| Item type | Current location | Collection | Call number | Status | Date due | Barcode | Item holds |
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Thesis
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UVAS Library Thesis Section | Veterinary Science | 1375,T (Browse shelf) | Available | 1375,T |
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Escherichia coli is normal inhabitant of all the animals and human beings. The Sorbitol
non fermenting E. coli strains were detected in milk, beef and fecal samples collected
from different areas of Lahore. White colored colonies of each positive sample on
Sorbitol MacConkey's Agar (SMA) contained gram negative, rods with round ends non
spore former Escherichia coli. Each of the isolates was sorbitol non fermenter, lactose
fermenter, indole positive, Methyl Red positive, Voges Prauskaur negative and citrate
negative. Each of the isolate was further characterized using polymerase chain reaction
(PCR) for the presence of shiga toxin 1 and shiga toxin 2 genes. Bacterial DNA was
extracted easily by boiling method when the isolates were grown on Sorbitol
MacConkey's agar at 37°C for 12-24 hours. The DNA was recovered when the culture
was boiled for 5-10 minutes. The isolated DNA when amplified using Stxland Stx2
specific primers showed that 68.5 percent samples were positive for Stxl and 54.2
percent for Stx2. The stx 1 and stx2 PCR products were subjected to sequencing. The
resulted sequences when aligned with the reference sequence through Basic local
alignment tool it showed that the shiga toxin 1 and shiga toxin 2 gene sequences are
conserved and showed high similarity in their nucleotide structure. Despite of having
high similarity in their nucleotide structure some haplotypes were also obtained showing
single nucleotide polymorphism. Phylogenetic analysis made among local isolates and
also with reported sequences from all over the world by using bioinformatics software to
see the genetic similarities and difference between them. The data produced showed
some highly conserved sequences and SNPs as well that will be quite useful for further
applications in diagnostics and biotechnology applications in future.
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