Difference between revisions of "Team:Tianjin/Note/6803"

Line 88: Line 88:
 
<article id="post-4252" class="post-4252 post type-post status-publish format-standard has-post-thumbnail hentry category-150 tag-174 tag-xinjiang tag-173">
 
<article id="post-4252" class="post-4252 post type-post status-publish format-standard has-post-thumbnail hentry category-150 tag-174 tag-xinjiang tag-173">
 
<header class="entry-header">
 
<header class="entry-header">
−
<h1 class="entry-title"><a href="https://www.camarts.cn/archives/4252.html" title="查看《火影忍者疾风传》中的全部作品" rel="bookmark">Week2(8/15/2016-8/21/2016)</a></h1>
+
<h1 class="entry-title">Week2(8/15/2016-8/21/2016)</h1>
 
</header><!-- .entry-header -->
 
</header><!-- .entry-header -->
  
Line 95: Line 95:
  
 
<a href="https://www.camarts.cn/archives/4252.html"><img class="aligncenter size-full wp-image-4256" src="http://static.camarts.cn/images/spinner-1600.gif"  alt="IMG_2956" width="800" height="533" srcset="images/img_1.jpg 800w, http://img.camarts.cn/2016/01/IMG_2956-150x100.jpg 150w, http://img.camarts.cn/2016/01/IMG_2956-300x200.jpg 300w, http://img.camarts.cn/2016/01/IMG_2956-768x512.jpg 768w, http://img.camarts.cn/2016/01/IMG_2956-450x300.jpg 450w" sizes="(max-width: 800px) 100vw, 800px" /><noscript><img class="aligncenter size-full wp-image-4256" src="http://img.camarts.cn/2016/01/IMG_2956.jpg?imageView/2/w/800/h/800/q/90" alt="IMG_2956" width="800" height="533" srcset="http://img.camarts.cn/2016/01/IMG_2956.jpg 800w, http://img.camarts.cn/2016/01/IMG_2956-150x100.jpg 150w, http://img.camarts.cn/2016/01/IMG_2956-300x200.jpg 300w, http://img.camarts.cn/2016/01/IMG_2956-768x512.jpg 768w, http://img.camarts.cn/2016/01/IMG_2956-450x300.jpg 450w" sizes="(max-width: 800px) 100vw, 800px" /></noscript></a></p>
 
<a href="https://www.camarts.cn/archives/4252.html"><img class="aligncenter size-full wp-image-4256" src="http://static.camarts.cn/images/spinner-1600.gif"  alt="IMG_2956" width="800" height="533" srcset="images/img_1.jpg 800w, http://img.camarts.cn/2016/01/IMG_2956-150x100.jpg 150w, http://img.camarts.cn/2016/01/IMG_2956-300x200.jpg 300w, http://img.camarts.cn/2016/01/IMG_2956-768x512.jpg 768w, http://img.camarts.cn/2016/01/IMG_2956-450x300.jpg 450w" sizes="(max-width: 800px) 100vw, 800px" /><noscript><img class="aligncenter size-full wp-image-4256" src="http://img.camarts.cn/2016/01/IMG_2956.jpg?imageView/2/w/800/h/800/q/90" alt="IMG_2956" width="800" height="533" srcset="http://img.camarts.cn/2016/01/IMG_2956.jpg 800w, http://img.camarts.cn/2016/01/IMG_2956-150x100.jpg 150w, http://img.camarts.cn/2016/01/IMG_2956-300x200.jpg 300w, http://img.camarts.cn/2016/01/IMG_2956-768x512.jpg 768w, http://img.camarts.cn/2016/01/IMG_2956-450x300.jpg 450w" sizes="(max-width: 800px) 100vw, 800px" /></noscript></a></p>
−
<p> <a href="https://www.camarts.cn/archives/4252.html" class="more-link">查看Team Tianjin全部实验 <span class="meta-nav">&rsaquo;</span></a></p>
 
−
</div><!-- .entry-content -->
 
−
 
−
<footer class="entry-meta">
 
−
       
 
−
<span class="sep">发表于 </span>2016 年 1 月 28 日<span class="sep"> | </span>
 
−
 
−
<span class="cat-links">
 
−
<span class="entry-utility-prep entry-utility-prep-cat-links">发表在</span> <a href="https://www.camarts.cn/archives/category/%e8%a5%bf%e5%8c%97%e8%a1%8c" rel="category tag">春野樱</a> </span>
 
−
<span class="sep"> | </span>
 
−
<span class="tag-links">
 
−
<span class="entry-utility-prep entry-utility-prep-tag-links">标签有</span> <a href="https://www.camarts.cn/archives/tag/%e5%b8%83%e5%b0%94%e6%b4%a5" rel="tag">春野樱</a>、<a href="https://www.camarts.cn/archives/tag/xinjiang" rel="tag">新疆</a>、<a href="https://www.camarts.cn/archives/tag/%e9%98%bf%e5%8b%92%e6%b3%b0" rel="tag">漩涡鸣人</a> </span>
 
−
 
−
<span class="sep"> | </span>
 
−
<span class="total-pictures"><a href="https://www.camarts.cn/archives/4252.html">5 张图片</a></span>
 
−
 
−
        </footer><!-- #entry-meta -->
 
 
          
 
          
 
<hr class="article">
 
<hr class="article">

Revision as of 12:34, 2 October 2016

TEAM TIANJIN

LOGO


Team Tianjin-Attribution

Notes

Week1(8/1/2016-8/7/2016)

  • Cultivation: 30µL mother liquid of Synechococcus sp. PCC 7942 (wild type)were cultivated in 10 mL BG-11-media at 30°C and 140rpm.

    • Week2(8/15/2016-8/21/2016)

  • Synechococcus sp PCC 7942 had no signals of life.

    • IMG_2956

    Week3(8/29/2016-9/4/2016)

  • pMV-G19
  • pMV-G15
  • Colonies were used to inoculate overnight cultures.
  • Plasmids were isolated using a miniprep kit.
    • Amplification of 19 and 15 with Q5 High-Fidelity DNA Polymerase out of E.coli
  • 19 amplification at 65.0°C with 19.rev/fwd primes.
    • PCR worked, positive control worked, no amplification of 19.
  • 15 amplification at 65.0°C with 15.rev/fwd primes.
    • PCR worked, positive control worked, no amplification of 15.
  • A product length of 900 bp was expected. The gel shows that most plasmids seem to be correct.
  • This result was confirmed by sequencing.
  • The fragments of 19 and 15 were purified with PCR Purification Kit.
    • Show More Ligation of 15 with 19
  • We add a single 3`-adenine overhang to each end of the fragment of 19 and then purified with DNA Purification Kit.
  • 19 was ligated into T vectors via TA clone and transformed into E.coli via heat shock.
  • A colony PCR was performed with five colonies.
    • Gel electrophoresis showed that 5 colonies were positive for insertion of 19. Two of these colonies containing 19 were used to inoculate overnight cultures.
  • 15 gene fragment was phosphorylated.
  • Plasmids containing T vector with 19(pT-19)were isolated using a miniprep kit.
  • Mono-restriction digest of pT-19 with stu I.
  • The enzyme-digested product was dephosphorylation.
  • Dephosphorylated plasmid and phosphorylated gene 15 were connected.
    • Ligation product was transformed into E.coli via heat shock.
  • A colony PCR was performed with twelve colonies.
    • Gel electrophoresis showed that 2 colonies were positive for connecting the plasmid and gene fragment. These two colonies were used to inoculate overnight cultures.
  • Plasmids with correct sequence of 19-15 were isolated using a miniprep kit.
  • Mono-restriction digest of pT-19-15 with Nru I.
  • The enzyme-digested product was dephosphorylation.
    • Show More IMG_2956

    查看Team Tianjin全部实验 ›

    Week4(9/5/2016-9/11/2016)

  • Colonies containing gene 13 were used to inoculate overnight cultures.
  • Plasmids were isolated using a miniprep kit.
    • Amplification of 13 with Q5 High-Fidelity DNA Polymerase out of E.coli
  • 13 amplification at 65.0°C with 13.rev/fwd primes
    • PCR worked, positive control worked, no amplification of 13
  • The fragments of 13 were purified with PCR Purification Kit.
  • 13 gene fragment was phosphorylated.
    • Show More Insertion of Ni promoter and ligation of 13-19-15
  • Ni inducible promoter was ligated into pCPC-3301 vector.
  • Phosphorylated 13 was ligated into pT-19-15 and transformed into E.coli via heat shock.
  • Single colonies were obtained by plating.
  • A colony PCR of Ni inducible promoter(pCPC-3301-Ni) was performed with 12 colonies.
    • Two of the successful ones were used to inoculate overnight cultures.
  • A colony PCR of pT-13-19-15 was performed with 7 colonies.
    • Two of the successful ones were used to inoculate overnight cultures.
  • Two kinds of plasmids were isolated using a miniprep kit.
    • Show More IMG_2956

    查看Team Tianjin全部实验 ›

    Week5(9/12/2016-9/18/2016)

  • Plasmids pT-13-19-15 were handed in for sequencing, which confirmed its correctness.
    • The sequencing results for both of them were error.
  • Colonies containing Ni inducible promoter were used to inoculate overnight cultures.
  • PCR was performed to check if the gene fragments were ligated correctly.
    • 13_ fwd and 15_rev on pT-13-19-15
    Gel electrophoresis showed that it failed.
  • Plasmids pCPC-3031-Ni were isolated using a miniprep kit.
  • Several PCRs were performed to check if the gene fragments were ligated correctly.
    • 1.13_fwd and 13_rev on pT-13-19-15
    2.19_fwd and 19_rev on pT-13-19-15
    3.15_fwd and 15_rev on pT-13-19-15
    4.13_fwd and 19_rev on pT-13-19-15
    5.19_fwd and 15_rev on pT-13-19-15
    6.13_ wd and 15_rev on pT-13-19-15
    The fourth and sixth ones were not successful.
  • Repetition: Phosphorylated 13 was ligated into pT-19-15 and transformed into E.coli via heat shock.
  • Repetition: Several PCRs were performed to check if the gene fragments were ligated correctly.
    • 1.13_fwd and 13_rev on pT-13-19-15
    2.13_fwd and 19_rev on pT-13-19-15
    The second one was failed.
    IMG_2956

    查看Team Tianjin全部实验 ›

    Week6(9/19/2016-9/25/2016)

  • Medium preparation :BG-11.
  • 19_ fwd and 15_rev were used to amplify 19-15.
    • Gel electrophoresis showed that amplification of fragments was successfull.
  • Ligated 13 and 19-15 via overlap PCR.
    • This PCR worked well and the products were extracted from the gel using a Agarose Gel DNA Extration Kit.
  • Restriction digest on pCPC-3031-Ni with Sac I.
  • The fragment 13-19-15 was ligated onto T vector and transformed into E.coli via heat shock.
  • A colony PCR of pT-13-19-15 was performed with 12 colonies.
    • Two of these colonies containing pT-13-19-15 were used to inoculate overnight cultures.
  • 13-19-15 gene fragment was phosphorylated.
  • Phosphorylated 13-19-15 was ligated into pCPC-3031-Ni and transformed into E.coli via heat shock.
  • Plasmids pT-13-19-15 were isolated using a miniprep kit.
  • A colony PCR of pCPC-3031-Ni-13-19-15 was performed with 12 colonies.
    • Three colonies were proved to be true. And two of them were used to inoculate overnight cultures.
    IMG_2956

    查看Team Tianjin全部实验 ›

    info_outline
    Notice: This page is currently under construction. Contents in this page are temporaory and will be modified several times before the final release.     — 2016 iGEM Team Tianjin