-
Notifications
You must be signed in to change notification settings - Fork 0
/
Copy pathBRED.html
509 lines (498 loc) · 37.1 KB
/
BRED.html
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
61
62
63
64
65
66
67
68
69
70
71
72
73
74
75
76
77
78
79
80
81
82
83
84
85
86
87
88
89
90
91
92
93
94
95
96
97
98
99
100
101
102
103
104
105
106
107
108
109
110
111
112
113
114
115
116
117
118
119
120
121
122
123
124
125
126
127
128
129
130
131
132
133
134
135
136
137
138
139
140
141
142
143
144
145
146
147
148
149
150
151
152
153
154
155
156
157
158
159
160
161
162
163
164
165
166
167
168
169
170
171
172
173
174
175
176
177
178
179
180
181
182
183
184
185
186
187
188
189
190
191
192
193
194
195
196
197
198
199
200
201
202
203
204
205
206
207
208
209
210
211
212
213
214
215
216
217
218
219
220
221
222
223
224
225
226
227
228
229
230
231
232
233
234
235
236
237
238
239
240
241
242
243
244
245
246
247
248
249
250
251
252
253
254
255
256
257
258
259
260
261
262
263
264
265
266
267
268
269
270
271
272
273
274
275
276
277
278
279
280
281
282
283
284
285
286
287
288
289
290
291
292
293
294
295
296
297
298
299
300
301
302
303
304
305
306
307
308
309
310
311
312
313
314
315
316
317
318
319
320
321
322
323
324
325
326
327
328
329
330
331
332
333
334
335
336
337
338
339
340
341
342
343
344
345
346
347
348
349
350
351
352
353
354
355
356
357
358
359
360
361
362
363
364
365
366
367
368
369
370
371
372
373
374
375
376
377
378
379
380
381
382
383
384
385
386
387
388
389
390
391
392
393
394
395
396
397
398
399
400
401
402
403
404
405
406
407
408
409
410
411
412
413
414
415
416
417
418
419
420
421
422
423
424
425
426
427
428
429
430
431
432
433
434
435
436
437
438
439
440
441
442
443
444
445
446
447
448
449
450
451
452
453
454
455
456
457
458
459
460
461
462
463
464
465
466
467
468
469
470
471
472
473
474
475
476
477
478
479
480
481
482
483
484
485
486
487
488
489
490
491
492
493
494
495
496
497
498
499
500
501
502
503
504
505
506
507
508
509
<!doctype html>
<html class="no-js" lang="en-US">
<head>
<meta charset="UTF-8">
<meta http-equiv="x-ua-compatible" content="ie=edge">
<meta name="viewport" content="width=device-width, initial-scale=1">
<title>BRED System</title>
<link rel="stylesheet" href="https://webify-13e95.kxcdn.com/demo/webify/architect/wp-content/cache/min/2/cedee2bfdeb161a102c92fd0d0c5247a.css"
data-minify="1" />
<meta name="robots" content="index, follow, max-snippet:-1, max-image-preview:large, max-video-preview:-1" />
<link rel="canonical" href="https://themebubble.com/demo/webify/architect/portfolio/greek-tree-tower/" />
<meta property="og:locale" content="en_US" />
<meta property="og:type" content="article" />
<meta property="og:title" content="Greek Tree Tower - Architect" />
<meta property="og:description" content="A digital product for everyone who take charge, make noise, and drop-kick the glass ceiling. The product team viewed the platform selection process as an opportunity to crystalize how they do performance management and approached the search thoughtfully. “We evaluated pretty much every performance management tool that exists.” Our role was to create the project […]"
/>
<meta property="og:url" content="https://themebubble.com/demo/webify/architect/portfolio/greek-tree-tower/" />
<meta property="og:site_name" content="Architect" />
<meta property="article:modified_time" content="2019-04-24T15:34:01+00:00" />
<meta property="og:image" content="https://webify-13e95.kxcdn.com/demo/webify/architect/wp-content/uploads/sites/2/2019/03/sandro-katalina-219649-unsplash-2.jpg"
/>
<meta property="og:image:width" content="1920" />
<meta property="og:image:height" content="1281" />
<meta name="twitter:card" content="summary_large_image" />
<meta name="twitter:label1" content="Written by">
<meta name="twitter:data1" content="David Beck">
<meta name="twitter:label2" content="Est. reading time">
<meta name="twitter:data2" content="1 minute">
<script type="application/ld+json" class="yoast-schema-graph">
{"@context":"https://schema.org","@graph":[{"@type":"WebSite","@id":"https://themebubble.com/demo/webify/architect/#website","url":"https://themebubble.com/demo/webify/architect/","name":"Architect","description":"Just another Webify Sites site","potentialAction":[{"@type":"SearchAction","target":"https://themebubble.com/demo/webify/architect/?s={search_term_string}","query-input":"required name=search_term_string"}],"inLanguage":"en-US"},{"@type":"ImageObject","@id":"https://themebubble.com/demo/webify/architect/portfolio/greek-tree-tower/#primaryimage","inLanguage":"en-US","url":"https://webify-13e95.kxcdn.com/demo/webify/architect/wp-content/uploads/sites/2/2019/03/sandro-katalina-219649-unsplash-2.jpg","width":1920,"height":1281},{"@type":"WebPage","@id":"https://themebubble.com/demo/webify/architect/portfolio/greek-tree-tower/#webpage","url":"https://themebubble.com/demo/webify/architect/portfolio/greek-tree-tower/","name":"Greek Tree Tower - Architect","isPartOf":{"@id":"https://themebubble.com/demo/webify/architect/#website"},"primaryImageOfPage":{"@id":"https://themebubble.com/demo/webify/architect/portfolio/greek-tree-tower/#primaryimage"},"datePublished":"2019-04-09T16:05:25+00:00","dateModified":"2019-04-24T15:34:01+00:00","inLanguage":"en-US","potentialAction":[{"@type":"ReadAction","target":["https://themebubble.com/demo/webify/architect/portfolio/greek-tree-tower/"]}]}]}
</script>
<link rel='dns-prefetch' href='//fonts.googleapis.com' />
<link rel='dns-prefetch' href='//cdn.jsdelivr.net' />
<link rel='dns-prefetch' href='//webify-13e95.kxcdn.com' />
<link rel="alternate" type="application/rss+xml" title="Architect » Feed" href="https://themebubble.com/demo/webify/architect/feed/"
/>
<link rel="alternate" type="application/rss+xml" title="Architect » Comments Feed" href="https://themebubble.com/demo/webify/architect/comments/feed/"
/>
<style type="text/css">
img.wp-smiley,
img.emoji {
display: inline !important;
border: none !important;
box-shadow: none !important;
height: 1em !important;
width: 1em !important;
margin: 0 .07em !important;
vertical-align: -0.1em !important;
background: none !important;
padding: 0 !important
}
</style>
<link rel='stylesheet' id='webify-fonts-css' href='https://fonts.googleapis.com/css?family=Roboto%3A300%2C400%2C500%2C700&subset=latin%2Clatin-ext&ver=1.0.0'
type='text/css' media='all' />
<link rel='stylesheet' id='webify-font-awesome-css' href='https://cdn.jsdelivr.net/npm/@fortawesome/[email protected]/css/all.min.css?ver=1.0.0'
type='text/css' media='all' />
<link rel='stylesheet' id='webify-font-awesome-v4-css' href='https://cdn.jsdelivr.net/npm/@fortawesome/[email protected]/css/v4-shims.min.css?ver=1.0.0'
type='text/css' media='all' />
<link rel='stylesheet' id='material-icon-css' href='https://fonts.googleapis.com/icon?family=Material+Icons%7CMaterial+Icons+Outlined&ver=1.0.0'
type='text/css' media='all' />
<style id='webify-style-inline-css' type='text/css'>
.tb-error-page {
background-image: url(https://webify-13e95.kxcdn.com/demo/webify/architect/wp-content/themes/webify/assets/img/error-bg.jpg)
}
.tb-page-header {
background-image: url(https://webify-13e95.kxcdn.com/demo/webify/architect/wp-content/themes/webify/assets/img/page-header-bg.png)
}
</style>
<script type='text/javascript' src='https://webify-13e95.kxcdn.com/demo/webify/architect/wp-content/cache/busting/2/demo/webify/architect/wp-includes/js/jquery/jquery-1.12.4-wp.js'
id='jquery-core-js'>
</script>
<link rel="https://api.w.org/" href="https://themebubble.com/demo/webify/architect/wp-json/" />
<link rel="EditURI" type="application/rsd+xml" title="RSD" href="https://themebubble.com/demo/webify/architect/xmlrpc.php?rsd"
/>
<link rel="wlwmanifest" type="application/wlwmanifest+xml" href="https://themebubble.com/demo/webify/architect/wp-includes/wlwmanifest.xml"
/>
<meta name="generator" content="WordPress 5.5.4" />
<link rel='shortlink' href='https://themebubble.com/demo/webify/architect/?p=367' />
<link rel="alternate" type="application/json+oembed" href="https://themebubble.com/demo/webify/architect/wp-json/oembed/1.0/embed?url=https%3A%2F%2Fthemebubble.com%2Fdemo%2Fwebify%2Farchitect%2Fportfolio%2Fgreek-tree-tower%2F"
/>
<link rel="alternate" type="text/xml+oembed" href="https://themebubble.com/demo/webify/architect/wp-json/oembed/1.0/embed?url=https%3A%2F%2Fthemebubble.com%2Fdemo%2Fwebify%2Farchitect%2Fportfolio%2Fgreek-tree-tower%2F&format=xml"
/>
<link rel="icon" href="logo_small_no_text.png"
sizes="32x32" />
<link rel="icon" href="logo_small_no_text.png"
sizes="192x192" />
<link rel="apple-touch-icon" href="logo_small_no_text.png"
/>
<meta name="msapplication-TileImage" content="logo_small_no_text.png"
/>
<style type="text/css">
.tb-site-footer,
.tb-site-footer.tb-style2 {
background-image: linear-gradient(#0c161c, #0c161c);
background-color: #0c161c
}
.tb-site-footer .tb-newsletter.tb-style8 button {
background-color: #62b0e2
}
.tb-site-footer .tb-newsletter.tb-style8 button:hover {
background-color: #91c3e2
}
</style>
<noscript>
<style id="rocket-lazyload-nojs-css">
.rll-youtube-player,
[data-lazy-src] {
display: none !important
}
</style>
</noscript>
</head>
<body class="portfolio-template-default single single-portfolio postid-367 wp-embed-responsive elementor-default elementor-kit-646 elementor-page elementor-page-367">
<div class="tb-preloader">
<div class="tb-preloader-in">
<img src="logo_crop_v.png" alt="loader-logo">
</div>
</div>
<header class="tb-site-header tb-default-width tb-style1 tb-sticky-header tb-solid-header tb-color1 tb-header-border1">
<div class="tb-main-header">
<div class="container">
<div class="tb-main-header-in">
<div class="tb-main-header-left">
<div class="tb-site-branding">
<a href="http://www.neuvae.com/index.html" class="tb-custom-logo-link ">
<img src="logo_no_text.png" alt="logo" class="tb-custom-logo">
</a>
</div>
</div>
<div class="tb-main-header-right">
<nav class="tb-main-nav tb-primary-nav">
<ul id="nav" class="tb-primary-nav-list">
<li id="menu-item-566" class="menu-item menu-item-type-post_type menu-item-object-page menu-item-home current-menu-item page_item page-item-5 current_page_item menu-item-566">
<a href="index.html">Home</a>
</li>
<li id="menu-item-523" class="menu-item menu-item-type-post_type menu-item-object-page menu-item-523">
<a href="http://www.neuvae.com/problem.html">Problem</a>
</li>
<li id="menu-item-523" class="menu-item menu-item-type-post_type menu-item-object-page menu-item-523">
<a href="http://www.neuvae.com/solution.html">Solution</a>
</li>
<li id="menu-item-558" class="menu-item menu-item-type-post_type menu-item-object-page menu-item-558">
<a href="http://www.neuvae.com/media.html">Media</a>
</li>
<li id="menu-item-558" class="menu-item menu-item-type-post_type menu-item-object-page menu-item-558">
<a href="http://www.neuvae.com/contact.html">Contact</a>
</li>
</ul>
</nav>
</div>
</div>
</div>
</div>
</header>
<div class="tb-content">
<div class="tb-post tb-style6">
<div href="#" class="tb-post-thumb tb-bg" style="background-image: url(https://www.mdpi.com/applsci/applsci-10-07322/article_deploy/html/images/applsci-10-07322-g001.png);"></div>
<div class="tb-post-info">
<div class="container">
<h2 class="tb-post-title tb-f48-lg tb-m0">BRED System</h2>
<div class="tb-catagory tb-style1">
</div></div></div></div><div class="empty-space marg-lg-b70 marg-sm-b60"></div>
<div class="container">
<div class="row">
<div class="col-lg-12">
<div class="tt-sticky-content">
<div class="tt-sticky-content-middle">
<div class="tt-sticky-content-in">
<div class="tb-left-padd-20">
<div class="tb-study-body">
<div data-elementor-type="wp-post" data-elementor-id="367" class="elementor elementor-367" data-elementor-settings="[]">
<div class="elementor-inner">
<div class="elementor-section-wrap">
<p>Engineering a phage to increase its host range is one of the fundamental barriers to widespread adoption of novel phage treatments. This is especially true for the nature of phages as for each strain of a species, there is one if not a few hundred different variants of the phage. Although this serves several benefits in directed, personalized and targeted medicine, the case to be made when it comes to treating infections often comes down to multiple bacteria at the root of the problem. That makes creating cocktails of tens of hundreds of different naturally occurring phages not just time consuming and expensive, but also ineffective at treating the disease. </p>
<br>
<p>This is where the idea of genetically modifying the target binding regions of a phage’s tail came to be, where we can just create 1 phage that could now expand its host-range through its targeting mechanism. Now, you can formulate not just custom but scalable phages that can treat all strains of E.coli, or perhaps both E.coli and TB and so on. The only problem is that the most accurate methods out there are CRISPR-Cas9 and other genetic insertion/deletion methods which are just as costly and hard to scale as traditional phage cocktails. </p>
<br>
<p>Thankfully, we can exploit the natural mechanisms of phages and bacteria for our own benefit: mutations. Over years, bacteria have developed advanced genetic responses to invaders and threats that can trigger mutations through its resistome. It’s what makes antibiotic resistance such a challenge, but this is also why phages are able to fight back. While antibiotics are static chemical compounds which do not evolve or change in response to AMR, phages are the biological competitor to bacteria and have, albeit different but effective, expansive genetic triggers that can allow them to mutate and circumvent the resistance mechanisms of bacteria. This not only makes resistance less of a factor for phage therapy because the rate at which phages mutate and change is faster than that of the bacteria, but it also presents an opportunity for the phages to directly counteract the resistance mechanisms that are mutated (i.e. phages can better attack efflux pumps).</p>
<br>
<p>Through phage mutation however, we can selectively force mutations in phages to attack multiple kinds of strains through homologous recombineering. The system comprises injecting DNA into specific points of the host’s genome or location. This can be used for altering the binding receptors of a phage to increase its range. However, traditional recombineering methods are ineffective at this because the phages cannot use selective markers such as drug-resistance genes for targeting the bacteria during viral lytic growth. Thanks to mutagenesis through homologous recombineering, this problem has been met but current systems are slow, unable to be used for all kinds of phages, and are not great at identifying useful mutations in the phages over several generations. </p>
<br>
<p>This is where BRED comes in: BRED or Bacteriophage Recombineering of Electroporated DNA is a highly efficient recombineering system that primarily uses electroporated phage DNA. Electroporation is when you apply an electrical field to an organism and make its cell membrane more permeable for insertion of DNA, drugs, and chemicals. In BRED, this means that mutants can easily be detected by PCR over the DNA (easily visible DNA) and it expedites the process significantly. BRED can be used to construct new, unmarked gene deletions, in-frame internal deletions, base substitutions, gene replacements with precision, and introduce gene tags for sequence identification.</p>
<br>
<br>
<br>
<section class="elementor-section elementor-top-section elementor-element elementor-element-2f1f65d elementor-section-boxed elementor-section-height-default elementor-section-height-default" data-id="2f1f65d" data-element_type="section">
<div class="elementor-container elementor-column-gap-extended">
<div class="elementor-row">
<div class="elementor-column elementor-col-50 elementor-top-column elementor-element elementor-element-84a92f5" data-id="84a92f5" data-element_type="column">
<div class="elementor-column-wrap elementor-element-populated">
<div class="elementor-widget-wrap">
<div class="elementor-element elementor-element-90f909a elementor-widget elementor-widget-image" data-id="90f909a" data-element_type="widget" data-widget_type="image.default">
<div class="elementor-widget-container">
<br>
<div class="elementor-image"> <img width="768" height="586" src="https://images.unsplash.com/photo-1576086213369-97a306d36557?ixid=MnwxMjA3fDB8MHxwaG90by1wYWdlfHx8fGVufDB8fHx8&ixlib=rb-1.2.1&auto=format&fit=crop&w=800&q=80" class="attachment-medium_large size-medium_large" alt="" loading="lazy" sizes="(max-width: 768px) 100vw, 768px" /></div>
</div>
</div>
</div>
</div>
</div>
<div class="elementor-column elementor-col-50 elementor-top-column elementor-element elementor-element-41df41c" data-id="41df41c" data-element_type="column">
<div class="elementor-column-wrap elementor-element-populated">
<div class="elementor-widget-wrap">
<div class="elementor-element elementor-element-6a6946b elementor-widget elementor-widget-webify-text-block-with-signature-widget" data-id="6a6946b" data-element_type="widget" data-widget_type="webify-text-block-with-signature-widget.default">
<div class="elementor-widget-container">
<div class="tb-vertical-middle">
<div class="tb-text-box tb-style1">
<div class="empty-space marg-lg-b15"></div>
<p>Some techniques mediated by recombination proteins were used for E.coli using the lambda phage Red proteins, EXO AND BETA -> they then exploited Rac prophage for mutant construction, meaning that lytic growth is low and the role of antibiotic resistance pressure cannot be effectively used for mutant selection. Another key benefit is that thanks to the co-electroplating of both the phage DNA and targeting substrates, it becomes easier to engineer the substrate and its insertion, and when the mutations are then recovered, you can get a high proportion of copies that possess these mutations </p>
<br>
<p> Some techniques mediated by recombination proteins were used for E.coli using the lambda phage Red proteins, EXO AND BETA -> they then exploited Rac prophage for mutant construction, meaning that lytic growth is low and the role of antibiotic resistance pressure cannot be effectively used for mutant selection. Another key benefit is that thanks to the co-electroplating of both the phage DNA and targeting substrates, it becomes easier to engineer the substrate and its insertion, and when the mutations are then recovered, you can get a high proportion of copies that possess these mutations.</p>
</div>
</div>
</div>
</div>
</div>
</div>
</div>
</div>
</div>
</section>
<section class="elementor-section elementor-top-section elementor-element elementor-element-4778445 elementor-section-boxed elementor-section-height-default elementor-section-height-default" data-id="4778445" data-element_type="section">
<div class="elementor-container elementor-column-gap-extended">
<div class="elementor-row">
<div class="elementor-column elementor-col-50 elementor-top-column elementor-element elementor-element-a85a349" data-id="a85a349" data-element_type="column">
<div class="elementor-column-wrap elementor-element-populated">
<div class="elementor-widget-wrap">
<div class="elementor-element elementor-element-f7ff5c3 elementor-widget elementor-widget-webify-section-heading-widget" data-id="f7ff5c3" data-element_type="widget" data-widget_type="webify-section-heading-widget.default">
<div class="elementor-widget-container">
<div class="tb-section-heading tb-style2">
<div class="empty-space marg-lg-b10"></div>
</div>
</div>
</div>
<div class="elementor-element elementor-element-ccdbfd2 elementor-widget elementor-widget-spacer" data-id="ccdbfd2" data-element_type="widget" data-widget_type="spacer.default">
<div class="elementor-widget-container">
<div class="elementor-spacer">
<div class="elementor-spacer-inner"></div>
</div>
</div>
</div>
<div class="elementor-element elementor-element-e5720fb elementor-widget elementor-widget-webify-tabs-widget" data-id="e5720fb" data-element_type="widget" data-widget_type="webify-tabs-widget.default">
<div class="elementor-widget-container">
<div class="tb-tabs tb-fade-tabs tb-style1">
<div class="empty-space marg-lg-b20"></div>
<div class="tab-content">
<div id="001-myself" class="tb-tab active ">
<div class="tb-f16-lg tb-line1-6 tb-tab-content tb-about-text">
<p>The recovery methods for these mutations are similar to most pipelines: the plaque that is formed through the phage replication and spread + lysis will be purified and analyzed against a wild-type (naturally occurring mutation) through PCR. When analyzing the selective mutant and wild type, you can quickly observe where the point mutations have occurred and depending on how easily the mutated types can be extracted, the more viable they will be for phage engineering. BRED can automatically expedite the process of identifying more useful and viable phage mutants as the process can recover 2-4x more pure mutations per plaque. </p>
<br>
<p>In fact, mutant-containing plaques can be recovered at an efficacy of 3.4-22.2%. After initial PCR screening of plaques, future purification is still needed to isolate homogenous phage mutants. Once the mutants are isolated, they will be then generated by infecting bacterial cells with the wild-type phage which would let the process to scale, following which extensive screening of recombinants will be required so that we can then identify the correct antibody for delivery using phage display. </p>
</div>
</div>
</div>
</div>
</div>
</div>
<div class="elementor-element elementor-element-a64e331 elementor-widget elementor-widget-spacer" data-id="a64e331" data-element_type="widget" data-widget_type="spacer.default">
<div class="elementor-widget-container">
<div class="elementor-spacer">
<div class="elementor-spacer-inner"></div>
</div>
</div>
</div>
</div>
</div>
</div>
<div class="elementor-column elementor-col-50 elementor-top-column elementor-element elementor-element-cf1d2e7" data-id="cf1d2e7" data-element_type="column">
<div class="elementor-column-wrap elementor-element-populated">
<div class="elementor-widget-wrap">
<div class="elementor-element elementor-element-eb23fee elementor-widget elementor-widget-image" data-id="eb23fee" data-element_type="widget" data-widget_type="image.default">
<div class="elementor-widget-container">
<br><br>
<div class="elementor-image"> <img width="862" height="936" src="https://images.unsplash.com/photo-1576086686350-2f5dba3ffeb3?ixid=MnwxMjA3fDB8MHxwaG90by1wYWdlfHx8fGVufDB8fHx8&ixlib=rb-1.2.1&auto=format&fit=crop&w=852&q=80" alt="" loading="lazy" sizes="(max-width: 862px) 100vw, 862px" /></div>
</div>
</div>
</div>
</div>
</div>
</div>
</div>
</section>
<br><br><br>
<div class="tb-text-box-btn text-center">
<a href="https://phagesdb.org/media/workflow/protocols/pdfs/BRED_Main_Protocol_PDF.pdf" target="_self" class="tb-btn tb-btn-primary tb-style3 tb-color2 tb-font-name">Learn More about BRED</a>
</div>
</div>
</div>
</div>
</div>
</div>
</div>
</div>
</div>
</div>
</div>
</div>
<div class="empty-space marg-lg-b100 marg-sm-b60"></div>
</div>
<footer class="tb-site-footer tb-style1 tb-dark-footer tb-111-bg tb-sticky-footer">
<hr>
<div class="empty-space marg-lg-b40 marg-sm-b40"></div>
<div class="container">
<div class="row">
<div class="col-lg-12">
<div class="tb-cta tb-style1">
<div class="tb-cta-left">
<h2 class="tb-f18-lg tb-font-name tb-white-c6 tb-m0">Want to learn more?</h2>
<div class="empty-space marg-lg-b0 marg-sm-b30"></div>
</div>
<div class="tb-cta-right"> <a href="mailto:[email protected]" class="tb-btn tb-style3 tb-cta-btn tb-color1"><span>Email us</span></a></div>
</div>
</div>
</div>
</div>
<div class="empty-space marg-lg-b40 marg-sm-b40"></div>
<hr>
<div class="empty-space marg-lg-b60 marg-sm-b60"></div>
<div class="container">
<div class="row">
<div class="col-md-3 col-sm-6">
<div id="nav_menu-1" class="widget tb-footer-item tb-footer_widget widget_nav_menu">
<h2 class="tb-footer-widget-title tb-font-name"><span>Resources</span></h2>
<div class="menu-footer-one-container">
<ul id="menu-footer-one" class="menu">
<li id="menu-item-566" class="menu-item menu-item-type-post_type menu-item-object-page menu-item-home current-menu-item page_item page-item-5 current_page_item menu-item-566">
<a href="http://www.neuvae.com/index.html">Home</a>
</li>
<li id="menu-item-558" class="menu-item menu-item-type-post_type menu-item-object-page menu-item-558">
<a href="http://www.neuvae.com/problem.html">Problem</a>
</li>
<li id="menu-item-546" class="menu-item menu-item-type-post_type menu-item-object-page menu-item-546">
<a href="http://www.neuvae.com/solution.html">Solution</a>
</li>
<li id="menu-item-558" class="menu-item menu-item-type-post_type menu-item-object-page menu-item-558">
<a href="http://www.neuvae.com/media.html">Media</a>
</li>
<li>
<a href="http://www.neuvae.com/contact.html" class="menu-item menu-item-type-post_type menu-item-object-page menu-item-545">Contact Us</a>
<li>
</ul>
</div>
</div>
</div>
<div class="col-md-3 col-sm-6">
<div id="nav_menu-2" class="widget tb-footer-item tb-footer_widget widget_nav_menu">
<h2 class="tb-footer-widget-title tb-font-name"><span>Media</span></h2>
<div class="menu-footer-two-container">
<ul id="menu-footer-two" class="menu">
<li id="menu-item-478" class="menu-item menu-item-type-custom menu-item-object-custom menu-item-478"><a href="https://youtu.be/bGm_M9Ww_xQ">Video</a></li>
<li id="menu-item-480" class="menu-item menu-item-type-custom menu-item-object-custom menu-item-480"><a href="https://mikaelhaji.medium.com/3089c670c8f7">Article</a></li>
<li id="menu-item-481" class="menu-item menu-item-type-custom menu-item-object-custom menu-item-481"><a href="1pager.pdf">1-pager</a></li>
</ul>
</div>
</div>
</div>
<div class="col-md-3 col-sm-6">
<div id="webify_contact_details-1" class="widget tb-footer-item tb-footer_widget tb-footer-address-widget">
<h2 class="tb-footer-widget-title tb-font-name"><span>Contact Details</span></h2>
<ul class="tb-mp0">
<li><i class="fa fa-map-marker"></i>176 Younge Street, Toronto</li>
<li><i class="fa fa-phone"></i>[email protected]</li>
</ul>
</div>
</div>
<div class="col-md-3 col-sm-6">
<div id="webify_newsletter-2" class="widget tb-footer-item tb-footer_widget tb-widget-newsletter">
<h2 class="tb-footer-widget-title tb-font-name"><span>Sign up for Newsletter</span></h2>
<form method="post" action="mailto:[email protected]" onsubmit="return newsletter_check(this)" class="tb-newsletter tb-style8"> <input type="email" name="ne" required="" placeholder="Enter Email Address"> <button type="submit" class="tb-newsletter-submit"><i class="fa fa-paper-plane"></i></button></form>
</div>
</div>
</div>
</div>
<div class="empty-space marg-lg-b55 marg-sm-b55"></div>
<hr>
<div class="tb-copyright tb-style1 tb-f13-lg text-center">
<div class="container">
<div class="row">
<div class="col-lg-12">© Neuvae 2021. All rights reserved.</div>
</div>
</div>
</div>
</footer>
<div id="tb-scrollup"><i class="fa fa-angle-up"></i></div>
<div class="tb-video-popup">
<div class="tb-video-popup-overlay"></div>
<div class="tb-video-popup-content">
<div class="tb-video-popup-layer"></div>
<div class="tb-video-popup-container">
<div class="tb-video-popup-align">
<div class="embed-responsive embed-responsive-16by9"> <iframe class="embed-responsive-item" src="about:blank"></iframe></div>
</div>
<div class="tb-video-popup-close"></div>
</div>
</div>
</div>
</footer>
<div id="tb-scrollup">
<i class="fa fa-angle-up"></i>
</div>
<div class="tb-video-popup">
<div class="tb-video-popup-overlay"></div>
<div class="tb-video-popup-content">
<div class="tb-video-popup-layer"></div>
<div class="tb-video-popup-container">
<div class="tb-video-popup-align">
<div class="embed-responsive embed-responsive-16by9">
<iframe class="embed-responsive-item" src="about:blank"></iframe>
</div>
</div>
<div class="tb-video-popup-close"></div>
</div>
</div>
</div>
<script type='text/javascript' id='wp-api-fetch-js-after'>
wp.apiFetch.use(wp.apiFetch.createRootURLMiddleware("https://themebubble.com/demo/webify/architect/wp-json/"));
wp.apiFetch.nonceMiddleware = wp.apiFetch.createNonceMiddleware("3e0b1b716b");
wp.apiFetch.use(wp.apiFetch.nonceMiddleware);
wp.apiFetch.use(wp.apiFetch.mediaUploadMiddleware);
wp.apiFetch.nonceEndpoint = "https://themebubble.com/demo/webify/architect/wp-admin/admin-ajax.php?action=rest-nonce";
</script>
<script type='text/javascript' id='contact-form-7-js-extra'>
/* <![CDATA[ */
var wpcf7 = {
"cached": "1"
};
/* ]]> */
</script>
<script type='text/javascript' id='webify-main-js-extra'>
/* <![CDATA[ */
var get = {
"ajaxurl": "https:\/\/themebubble.com\/demo\/webify\/architect\/wp-admin\/admin-ajax.php",
"siteurl": "https:\/\/webify-13e95.kxcdn.com\/demo\/webify\/architect\/wp-content\/themes\/webify",
"nonce": "3554bd5532"
};
/* ]]> */
</script>
<script type='text/javascript' id='elementor-frontend-js-before'>
var elementorFrontendConfig = {
"environmentMode": {
"edit": false,
"wpPreview": false
},
"i18n": {
"shareOnFacebook": "Share on Facebook",
"shareOnTwitter": "Share on Twitter",
"pinIt": "Pin it",
"download": "Download",
"downloadImage": "Download image",
"fullscreen": "Fullscreen",
"zoom": "Zoom",
"share": "Share",
"playVideo": "Play Video",
"previous": "Previous",
"next": "Next",
"close": "Close"
},
"is_rtl": false,
"breakpoints": {
"xs": 0,
"sm": 480,
"md": 768,
"lg": 1025,
"xl": 1440,
"xxl": 1600
},
"version": "3.0.8",
"is_static": false,
"legacyMode": {
"elementWrappers": true
},
"urls": {
"assets": "https:\/\/themebubble.com\/demo\/webify\/architect\/wp-content\/plugins\/elementor\/assets\/"
},
"settings": {
"page": [],
"editorPreferences": []
},
"kit": {
"global_image_lightbox": "yes",
"lightbox_enable_counter": "yes",
"lightbox_enable_fullscreen": "yes",
"lightbox_enable_zoom": "yes",
"lightbox_enable_share": "yes",
"lightbox_title_src": "title",
"lightbox_description_src": "description"
},
"post": {
"id": 367,
"title": "Greek%20Tree%20Tower%20-%20Architect",
"excerpt": "\n\t\t\t\t\t\t",
"featuredImage": "https:\/\/webify-13e95.kxcdn.com\/demo\/webify\/architect\/wp-content\/uploads\/sites\/2\/2019\/03\/sandro-katalina-219649-unsplash-2.jpg"
}
};
</script>
<script src="https://webify-13e95.kxcdn.com/demo/webify/architect/wp-content/cache/min/2/e67e8431468a06ddb528547c0157238c.js" data-minify="1" defer></script>
</body></html>
<!-- This website is like a Rocket, isn't it? Performance optimized by WP Rocket. Learn more: https://wp-rocket.me - Debug: cached@1619386191 -->