p47 PCR on samples from flies injected with P4 DiNV to look at viral load

Rob wanted me to look at p47 banding on the flies I injected with P4 DiNV then sampled at 0, 1, 3, and 5 days post infection to measure viral load with qPCR. Flies were injected here, and run through qPCR here

These samples were 1:10 diluted for qPCR, so I used the diluted DNA for regular PCR as well, but increased the volume of DNA to 3ul in the reactions.

Information on the primers and their programs can be found here

The PCR process followed this general protocol.

Two sets of 96 reactions were run:

  • 500ul GoTaq
  • 25ul F p47 primer
  • 25ul R p47 primer
  • 150ul molecular grade water

Reactions were then run on a 1% gel for 40 min at 90V and scored for band presence, information found here:

Sample dilution day p47
29 0.01 FFU day0 no
116 0.01 FFU day3 no
31 0.01 FFU day0 no
118 0.01 FFU day3 yes
33 0.01 FFU day0 no
120 0.01 FFU day3 no
35 0.01 FFU day0 no
122 0.01 FFU day3 no
21 0.01 FFU day0 no
107 0.01 FFU day3 no
23 0.01 FFU day0 no
109 0.01 FFU day3 no
25 0.01 FFU day0 no
111 0.01 FFU day3 yes
27 0.01 FFU day0 no
113 0.01 FFU day3 yes
11 0.1 FFU day0 no
135 0.1 FFU day3 yes
13 0.1 FFU day0 no
137 0.1 FFU day3 yes
15 0.1 FFU day0 no
139 0.1 FFU day3 yes
17 0.1 FFU day0 no
141 0.1 FFU day3 yes
1 0.1 FFU day0 no
126 0.1 FFU day3 yes
3 0.1 FFU day0 no
128 0.1 FFU day3 yes
4 0.1 FFU day0 no
130 0.1 FFU day3 yes
5 0.1 FFU day0 no
132 0.1 FFU day3 yes
45 3 FFU day0 no
153 3 FFU day3 yes
47 3 FFU day0 no
155 3 FFU day3 yes
49 3 FFU day0 no
157 3 FFU day3 yes
51 3 FFU day0 no
159 3 FFU day3 yes
37 3 FFU day0 no
144 3 FFU day3 yes
39 3 FFU day0 no
146 3 FFU day3 yes
41 3 FFU day0 no
148 3 FFU day3 yes
43 3 FFU day0 no
150 3 FFU day3 yes
63 0.01 FFU day1 no
169 0.01 FFU day5 yes
65 0.01 FFU day1 no
171 0.01 FFU day5 no
67 0.01 FFU day1 no
173 0.01 FFU day5 no
69 0.01 FFU day1 no
175 0.01 FFU day5 no
53 0.01 FFU day1 no
161 0.01 FFU day5 no
55 0.01 FFU day1 no
163 0.01 FFU day5 no
57 0.01 FFU day1 no
165 0.01 FFU day5 yes
59 0.01 FFU day1 no
167 0.01 FFU day5 no
82 0.1 FFU day1 no
189 0.1 FFU day5 yes
84 0.1 FFU day1 no
191 0.1 FFU day5 yes
86 0.1 FFU day1 no
193 0.1 FFU day5 yes
88 0.1 FFU day1 no
195 0.1 FFU day5 yes
72 0.1 FFU day1 no
178 0.1 FFU day5 yes
74 0.1 FFU day1 no
180 0.1 FFU day5 yes
76 0.1 FFU day1 no
182 0.1 FFU day5 yes
78 0.1 FFU day1 no
184 0.1 FFU day5 yes
99 3 FFU day1 yes
209 3 FFU day5 yes
101 3 FFU day1 yes
210 3 FFU day5 yes
103 3 FFU day1 yes
211 3 FFU day5 yes
105 3 FFU day1 yes
212 3 FFU day5 yes
91 3 FFU day1 yes
199 3 FFU day5 yes
93 3 FFU day1 yes
201 3 FFU day5 yes
95 3 FFU day1 yes
203 3 FFU day5 yes
97 3 FFU day1 yes
205 3 FFU day5 yes
30 0.01 FFU day0 no
117 0.01 FFU day3 no
32 0.01 FFU day0 no
119 0.01 FFU day3 yes
34 0.01 FFU day0 no
121 0.01 FFU day3 no
36 0.01 FFU day0 no
123 0.01 FFU day3 no
22 0.01 FFU day0 no
108 0.01 FFU day3 no
24 0.01 FFU day0 no
110 0.01 FFU day3 yes
26 0.01 FFU day0 no
112 0.01 FFU day3 no
28 0.01 FFU day0 no
114 0.01 FFU day3 no
12 0.1 FFU day0 no
136 0.1 FFU day3 yes
14 0.1 FFU day0 no
138 0.1 FFU day3 yes
16 0.1 FFU day0 no
140 0.1 FFU day3 yes
18 0.1 FFU day0 no
142 0.1 FFU day3 no
2 0.1 FFU day0 no
127 0.1 FFU day3 yes
6 0.1 FFU day0 no
129 0.1 FFU day3 yes
7 0.1 FFU day0 no
131 0.1 FFU day3 yes
8 0.1 FFU day0 no
133 0.1 FFU day3 yes
46 3 FFU day0 no
154 3 FFU day3 yes
48 3 FFU day0 no
156 3 FFU day3 yes
50 3 FFU day0 no
158 3 FFU day3 yes
52 3 FFU day0 no
160 3 FFU day3 yes
38 3 FFU day0 no
145 3 FFU day3 yes
40 3 FFU day0 no
147 3 FFU day3 yes
42 3 FFU day0 no
149 3 FFU day3 yes
44 3 FFU day0 no
151 3 FFU day3 yes
64 0.01 FFU day1 no
170 0.01 FFU day5 no
66 0.01 FFU day1 no
172 0.01 FFU day5 no
68 0.01 FFU day1 no
174 0.01 FFU day5 no
70 0.01 FFU day1 no
176 0.01 FFU day5 no
54 0.01 FFU day1 no
162 0.01 FFU day5 no
56 0.01 FFU day1 no
164 0.01 FFU day5 no
58 0.01 FFU day1 no
166 0.01 FFU day5 no
60 0.01 FFU day1 no
168 0.01 FFU day5 no
83 0.1 FFU day1 no
190 0.1 FFU day5 yes
85 0.1 FFU day1 no
192 0.1 FFU day5 yes
87 0.1 FFU day1 no
194 0.1 FFU day5 yes
89 0.1 FFU day1 yes
196 0.1 FFU day5 yes
73 0.1 FFU day1 no
179 0.1 FFU day5 yes
75 0.1 FFU day1 yes
181 0.1 FFU day5 yes
77 0.1 FFU day1 no
183 0.1 FFU day5 yes
79 0.1 FFU day1 yes
185 0.1 FFU day5 yes
100 3 FFU day1 yes
213 3 FFU day5 yes
102 3 FFU day1 yes
214 3 FFU day5 yes
104 3 FFU day1 yes
215 3 FFU day5 yes
106 3 FFU day1 yes
216 3 FFU day5 yes
92 3 FFU day1 yes
200 3 FFU day5 yes
94 3 FFU day1 yes
202 3 FFU day5 yes
96 3 FFU day1 yes
204 3 FFU day5 yes
98 3 FFU day1 yes
206 3 FFU day5 yes