Difference between revisions of "Part:BBa K2365511"
 
(17 intermediate revisions by 3 users not shown)
Line 17: Line 17:
 
<partinfo>BBa_K2365511 parameters</partinfo>
 
<partinfo>BBa_K2365511 parameters</partinfo>
 
<!-- -->
 
<!-- -->
 +
====3-dimension structure====
 +
CecropinA (CA)is a naturally occurring, linear, cationic, 37-residue antimicrobial peptide. It has inhibitory effect on Aspergillus and Fusarium. The possible mechanism of killing bacteria is make the cell to form holes. This is the three-dimensional structure. It has the alpha helix and beta folding structure.
 
[[File:Model1.gif|300px]]
 
[[File:Model1.gif|300px]]
 
[[File:Model2.gif|300px]]
 
[[File:Model2.gif|300px]]
−
[[File:Model2.gif|300px]]  
+
[[File:Model3.gif|300px]]
 +
 
 +
====PCR and SDS-PAGE====
 +
We did PCR and agarose gel electrophoresis to test and verify cecropinA. And then we did Tris tricine-SDS-PAGE to confirm if he successfully expressed. It’s has 177 bases and the kDa is about 5.The results show cecropinA was constructed and expressed successful.
 +
 
 +
 
 
[[File:Cerope A 1.jpg|300px]]
 
[[File:Cerope A 1.jpg|300px]]
 
[[File:Cecropin A NAU-08.jpeg|300px]]
 
[[File:Cecropin A NAU-08.jpeg|300px]]
 +
====Zone of inhibition====
 +
In order to verify the function of cecropinA, after it Successful expression, we did the bacteriostatic test, the results show chemical pesticide (positive) works very well, cecropinA also has bacteriostatic effect  but can’t compare with it. It achieved lethality for Fusarium graminearum and spores at 5ug/ml. With the increase of concentration, the inhibitory effect is better.
 
[[File:Cecropin A zone.jpeg|500px|center]]
 
[[File:Cecropin A zone.jpeg|500px|center]]
 +
====OD 595nm====
 +
We design another experiment to detect the function  of cecropinA. There were five samples, including negative control,1ug/ml,10 ug/ml,50 ug/ml,100 ug/ml. after 24 hours, the value of OD595 showed with the increase of concentration of cecropinA, the number of Fusarium graminearum conidia decreased, it’s LD50 is about 10 ug/ml, which has a  good antibacterial effect.
 +
====MASS spectrometry report and HPLC analysis of Cecropin A====
 
[[File:Cecropin_A_595nm.png|600px|left]]
 
[[File:Cecropin_A_595nm.png|600px|left]]
 +
[[File:Cecropin A mass 11.png|600px|center]]
 +
[[File:HPLC report CC.png|600px|center]]
 +
The MASS spectrometry report and HPLC analysis of Cecropin A,  from the graph we can get the weight measured by MASS spectrometry is 4003.78U and the amino acid sequence is Lys-Try-Lys-leu-phe-Lys-Lys-ile-Glu-Lys-val-Gly-Gln-Asn-Ile-Arg-Asp-Gly-Ile-Ile-Lys-Ala-Gly-pro-Ala-val-Ala-val-val-Gly-Gln-Ala-Thr-Gln-Ile-Ala-Lys-NH2

Latest revision as of 20:44, 1 November 2017


cecropinA

BBa_K2365511 encodes Cecropin A ,a naturally occurring,linear,cationic,antimicrobial peptide,it can inhibit the growth of Fusarium graminearum.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 14
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 106
    Illegal AgeI site found at 144
  • 1000
    COMPATIBLE WITH RFC[1000]


3-dimension structure

CecropinA (CA)is a naturally occurring, linear, cationic, 37-residue antimicrobial peptide. It has inhibitory effect on Aspergillus and Fusarium. The possible mechanism of killing bacteria is make the cell to form holes. This is the three-dimensional structure. It has the alpha helix and beta folding structure. Model1.gif Model2.gif Model3.gif

PCR and SDS-PAGE

We did PCR and agarose gel electrophoresis to test and verify cecropinA. And then we did Tris tricine-SDS-PAGE to confirm if he successfully expressed. It’s has 177 bases and the kDa is about 5.The results show cecropinA was constructed and expressed successful.


Cerope A 1.jpg Cecropin A NAU-08.jpeg

Zone of inhibition

In order to verify the function of cecropinA, after it Successful expression, we did the bacteriostatic test, the results show chemical pesticide (positive) works very well, cecropinA also has bacteriostatic effect but can’t compare with it. It achieved lethality for Fusarium graminearum and spores at 5ug/ml. With the increase of concentration, the inhibitory effect is better.

Cecropin A zone.jpeg

OD 595nm

We design another experiment to detect the function of cecropinA. There were five samples, including negative control,1ug/ml,10 ug/ml,50 ug/ml,100 ug/ml. after 24 hours, the value of OD595 showed with the increase of concentration of cecropinA, the number of Fusarium graminearum conidia decreased, it’s LD50 is about 10 ug/ml, which has a good antibacterial effect.

MASS spectrometry report and HPLC analysis of Cecropin A

Cecropin A 595nm.png
Cecropin A mass 11.png
HPLC report CC.png

The MASS spectrometry report and HPLC analysis of Cecropin A, from the graph we can get the weight measured by MASS spectrometry is 4003.78U and the amino acid sequence is Lys-Try-Lys-leu-phe-Lys-Lys-ile-Glu-Lys-val-Gly-Gln-Asn-Ile-Arg-Asp-Gly-Ile-Ile-Lys-Ala-Gly-pro-Ala-val-Ala-val-val-Gly-Gln-Ala-Thr-Gln-Ile-Ala-Lys-NH2