How to be a Breast Cancer Detective.

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Briefly describe the lesson here.  
Briefly describe the lesson here.  
-
The lesson begins with interpreting familial pedigree charts and continues with steps in the identification of the genes responsible for some breast cancers. It continues with gene analysis and identification of the mutations through bioinformatics techniques.
+
The lesson begins with interpreting familial pedigree charts and continues with steps in the identification of the genes responsible for some breast cancers. It continues with searching for the BRCA1 gene and analysis and identification of the coding sequences through bioinformatics techniques.
--------------------------------------------------------------------------------
--------------------------------------------------------------------------------
-
'''Goals and Objectives'''  
+
'''Goals and Objectives'''
 +
1) understand the process by which genes of parents are transferred to their offspring.  
1) understand the process by which genes of parents are transferred to their offspring.  
-
2) understand the difference between a dominant and recessive trait and understand how the presence of a trait may effect the physical characteristics of an individual.   
+
 
 +
2) understand the difference between a dominant and recessive trait and understand how the presence of a trait may effect the physical characteristics of an individual.  
 +
   
3) read and/or construct a pedigree chart mapping a specific trait in a family.  
3) read and/or construct a pedigree chart mapping a specific trait in a family.  
-
4) determine the probability of a certain phenotype being expressed in an individual.
 
-
5) learn techniques of searching for specific genes in databases.
 
-
6) using bioinformatics technology to compare normal and mutated genes
 
-
'''Common Misconceptions'''
+
4) determine the probability of a certain phenotype being expressed in a pedigree chart
-
Describe any common misconceptions this lesson may address
+
5) Understand the uses of Biotechnology and Bioinformatics in understanding cancer
 +
6) Learn to retrieve information from a Bioinformatics source.
 +
'''Common Misconceptions'''
 +
 +
 +
 +
Students may not understand the importance and relevance of molecular
 +
genetics to research on today's world. Many students are not aware of the world of Bioinformatics.
 +
 +
--------------------------------------------------------------------------------
--------------------------------------------------------------------------------
'''The Lesson'''  
'''The Lesson'''  
-
Preparation Before class:''' (materials, handouts etc.)'''
+
Preparation Before class
-
Part 1. Introduction to Pedigree charts (handout)Highlighters, pens, pencils, books, chalkboard, and 1 computer with LCD projector and sound system
+
 
 +
Part 1. PowerPoint to accompany lesson, Introduction to Pedigree charts (handout),Highlighters, pens, pencils, books, chalkboard, and 1 computer with LCD projector and sound system
 +
 
Part 2: Computer classroom; one computer per student, LCD projector,sound system  
Part 2: Computer classroom; one computer per student, LCD projector,sound system  
'''During class'''
'''During class'''
 +
 +
'''Time required'''
 +
 +
Part 1- 45 minutes
Part 1: Predicting the Breast cancer gene through Pedigree charts
Part 1: Predicting the Breast cancer gene through Pedigree charts
 +
         Practice drawing a pedigree chart
         Practice drawing a pedigree chart
          
          
Begin by using PowerPoint presentation which contains links to other resources listed below:
Begin by using PowerPoint presentation which contains links to other resources listed below:
 +
Mary Claire King: Finding brca1 and 2 by pedigree- http://www.dnai.org/media/a/king296¬04.swf
Mary Claire King: Finding brca1 and 2 by pedigree- http://www.dnai.org/media/a/king296¬04.swf
 +
Mark Skolnick: Finding the breast cancer gene brca1-http://www.dnai.org/media/a/skolnick298_06.swf
Mark Skolnick: Finding the breast cancer gene brca1-http://www.dnai.org/media/a/skolnick298_06.swf
 +
Mary Claire king: Can women be tested for breast cancer? http://www.dnaiorg/media/a/king295_08.swf
Mary Claire king: Can women be tested for breast cancer? http://www.dnaiorg/media/a/king295_08.swf
-
Part 2: Discussion of Biotechnology and bioinformatics based on reading [[Image:biotechnology.doc]]
+
 
 +
'''Time required'''
 +
 
 +
Part 2 -2 45 minute periods
 +
 
 +
How do mutations form?
 +
View film clip http://www.insidecancer.org/?s=A,1,7
 +
 
 +
1. Reading and Discussion of Biotechnology and Bioinformatics based on the reading below:
       Biotechnology’s Relation to Cancer Research  
       Biotechnology’s Relation to Cancer Research  
 +
Biotechnology has applied many of the techniques of cell biology,
Biotechnology has applied many of the techniques of cell biology,
molecular biology, and protein chemistry to cancer research and treatment over
molecular biology, and protein chemistry to cancer research and treatment over
Line 55: Line 83:
cascade of cancer events. Computer modeling allows proteins to be considered
cascade of cancer events. Computer modeling allows proteins to be considered
in a variety of new lights, including indentifying potential docking clefts for
in a variety of new lights, including indentifying potential docking clefts for
-
the introduction of small molecules or proteins that might interfere with cancer causing
+
the introduction of small molecules or proteins that might interfere with cancer causing protein functions. Bioinformatics and combinatorial chemistry allow
-
protein functions. Bioinformatics and combinatorial chemistry allow
+
scientists to sort through molecules (both natural and synthetic) that
scientists to sort through molecules (both natural and synthetic) that
interfere with cancer’s progress in a variety of ways. In addition, the ability
interfere with cancer’s progress in a variety of ways. In addition, the ability
Line 64: Line 91:
patterns in those measurements to help better understand how cancer cells
patterns in those measurements to help better understand how cancer cells
behave and how they react to different treatments.
behave and how they react to different treatments.
-
Fighting Cancer
+
(from Biotechnology and You,vol.11,issue 1)
-
        Finding the Brca 1 gene using bioinformatics
+
2. Finding the Brca 1 gene using bioinformatics
        
        
-
 
-
 
-
'''Time required'''
 
-
 
-
Part 1- 45 minutes
 
-
Part 2 -2 45 minute periods
 
'''Student Handouts for the Lesson Plan'''  
'''Student Handouts for the Lesson Plan'''  
-
Describe any handouts and provide links to documents that include the handouts. Remember to upload the handouts to the wiki before linking to them.
+
Instruction sheet on steps in finding genes and using the NCBI website
'''Alternative Assessments'''
'''Alternative Assessments'''
-
Describe any alternative activities or assessments you may have developed.  
+
Poster of Breast Cancer Pedigree with correct key
 +
 
 +
Bioinformatics project using tools and websites found in this lesson.
 +
 
'''Suggestions for Extended Learning'''
'''Suggestions for Extended Learning'''
Line 87: Line 111:
• Webster K Cavenee and Raymond L. White, “The Genetic Basis
• Webster K Cavenee and Raymond L. White, “The Genetic Basis
of Cancer,” Scientific American, March 1995.
of Cancer,” Scientific American, March 1995.
-
• Robert Cooke, Dr. Folkman’s War: Angiogenesis and the Struggle to
+
 
-
Defeat Cancer
+
-
• Jerome Groopman, “The Thirty Years’ War,” The New Yorker (June 4, 2001)
+
• Matt Ridley, Chapter 17 in Genome: Autobiography of a Species in 23 Chapters
• Matt Ridley, Chapter 17 in Genome: Autobiography of a Species in 23 Chapters
 +
• Robert Weinberg, One Renegade Cell
• Robert Weinberg, One Renegade Cell
-
• Lisa Yount, Cancer
+
 
• Time and Newsweek cover articles on cancer topics
• Time and Newsweek cover articles on cancer topics
 +
• American Cancer Society: http://www.cancer.org
• American Cancer Society: http://www.cancer.org
 +
• National Cancer Institute: http://www.nci.nih.gov
• National Cancer Institute: http://www.nci.nih.gov
 +
• National Childhood Cancer Foundation: http://www.nccf.org
• National Childhood Cancer Foundation: http://www.nccf.org
 +
• Oncolink: http://www.oncolink.upenn.edu
• Oncolink: http://www.oncolink.upenn.edu
 +
• NOVA’s program on Judah Folkman, Cancer Warrior: http://pbs.org/wgbh/
• NOVA’s program on Judah Folkman, Cancer Warrior: http://pbs.org/wgbh/
nova/cancer
nova/cancer
Line 104: Line 132:
'''Glossary'''  
'''Glossary'''  
 +
Allele—an alternate form of a gene.  
Allele—an alternate form of a gene.  
 +
Chromosome—structure in the cell nucleus that stores and transmits genetic information.  
Chromosome—structure in the cell nucleus that stores and transmits genetic information.  
 +
Gene—unit of heredity.  
Gene—unit of heredity.  
 +
Genotype—Allelic status of an organism for a genetic trait.  
Genotype—Allelic status of an organism for a genetic trait.  
 +
Heterozygous—having different alleles of a gene.  
Heterozygous—having different alleles of a gene.  
 +
Homozygous—having indistinguishable alleles of a gene.  
Homozygous—having indistinguishable alleles of a gene.  
-
Pedigree—diagram showing the expression of a specific characteristic and the biological relationships among members of a family, often of several generations.  
+
 
 +
Pedigree—diagram showing the expression of a specific characteristic and the biological relationships among members of a family, often of several generations.
 +
Phenotype—the observable organism, the expression of a genetic trait.  
Phenotype—the observable organism, the expression of a genetic trait.  
-
Bioinformatics-
 
-
NCBI-
 
-
ClustalW-
 
-
Brca 1 and 2-genes when mutated may cause breast cancer in humans
 
 +
Bioinformatics- the science of using databases to enter or research genetic information on the WWW.
 +
NCBI-National Center for Biotechnology Information
 +
 +
Biotechnology-the scientific manipulation of living organisms, especially at the molecular genetic level, to produce useful products.
 +
 +
CDS-Coding sequence
Line 123: Line 161:
'''Education Standards'''  
'''Education Standards'''  
-
Please align your lesson plan with any education standards that apply.  
+
• Living systems also have different levels of organization--for example, cells, tissues, organs, organisms, populations, and communities. Within these systems, interactions between components occur. Further, systems at different levels of organization can manifest different properties and functions.
 +
 
 +
• Models are tentative schemes or structures that correspond to real objects, events, or classes of events, and that have explanatory power. Models help scientists and engineers understand how things work. Models take many forms, including physical objects, plans, mental constructs, mathematical equations, and computer simulations.
 +
 
 +
• Cells have particular structures that underlie their functions. Every cell is surrounded by a membrane that separates it from the outside world. Inside the cell is a concentrated mixture of thousands of different molecules which form a variety of specialized structures that carry out such cell functions as energy production, transport of molecules, waste disposal, synthesis of new molecules, and the storage of genetic material.
 +
 
 +
• Cells store and use information to guide their functions. The genetic information stored in DNA is used to direct the synthesis of the thousands of proteins that each cell requires.
 +
 
 +
• In all organisms, the instructions for specifying the characteristics of the organism are carried in DNA, a large polymer formed from subunits of four kinds (A, G, C, and T). The chemical and structural properties of DNA explain how the genetic information that underlies heredity is both encoded in genes (as a string of molecular "letters") and replicated (by a templating mechanism). Each DNA molecule in a cell forms a single chromosome.
 +
 
 +
• Recognize and analyze alternative explanations and models, reviewing current scientific understanding.
'''Teacher Answer Key'''  
'''Teacher Answer Key'''  
 +
Part 2: Bioinformatics  Brca 1 gene:
Part 2: Bioinformatics  Brca 1 gene:
Questions
Questions
 +
1.What is a locus?  A position in the genome where a particular gene is found
1.What is a locus?  A position in the genome where a particular gene is found
 +
2.How many “bp” base pairs are found in this locus? 3759 base pairs
2.How many “bp” base pairs are found in this locus? 3759 base pairs
 +
3.Where is the gene found? mRNA Homo Sapiens
3.Where is the gene found? mRNA Homo Sapiens
 +
4.What does the gene encode for?  
4.What does the gene encode for?  
 +
             This gene encodes a nuclear phosphoprotein that plays a
             This gene encodes a nuclear phosphoprotein that plays a
             role in maintaining genomic stability and acts as a tumor
             role in maintaining genomic stability and acts as a tumor
Line 138: Line 192:
             suppressors, DNA damage sensors, and signal transducers to form a
             suppressors, DNA damage sensors, and signal transducers to form a
             large multi-subunit protein complex known as BASC for
             large multi-subunit protein complex known as BASC for
-
             BRCA1-associated genome surveillance complex. This gene product
+
             BRCA1-associated genome surveillance complex.  
-
            associates with RNA polymerase II, and through the C-terminal
+
 
-
            domain, also interacts with histone deacetylase complex. This
+
CDS of Human Breast cancer gene
-
            protein thus plays a role in transcription, DNA repair of
+
                                tggatttat ctgctcttcg cgttgaagaa gtacaaaatg
-
            double-stranded breaks, and recombination. Mutations in this gene
+
      241 tcattaatgc tatgcagaaa atcttagagt gtcccatctg tctggagttg atcaaggaac
-
            are responsible for approximately 40% of inherited breast cancers
+
      301 ctgtctccac aaagtgtgac cacatatttt gcaaattttg catgctgaaa cttctcaacc
-
            and more than 80% of inherited breast and ovarian cancers.
+
      361 agaagaaagg gccttcacag tgtcctttat gtaagaatga tataaccaaa aggagcctac
-
            Alternative splicing plays a role in modulating the subcellular
+
      421 aagaaagtac gagatttagt caacttgttg aagagctatt gaaaatcatt tgtgcttttc
-
            localization and physiological function of this gene. Many
+
      481 agcttgacac aggtttggag tatgcaaaca gctataattt tgcaaaaaag gaaaataact
-
            alternatively spliced transcript variants have been described for
+
      541 ctcctgaaca tctaaaagat gaagtttcta tcatccaaag tatgggctac agaaaccgtg
-
            this gene but only some have had their full-length natures
+
      601 ccaaaagact tctacagagt gaacccgaaa atccttcctt gcaggaaacc agtctcagtg
-
            identified.
+
      661 tccaactctc taaccttgga actgtgagaa ctctgaggac aaagcagcgg atacaacctc
 +
      721 aaaagacgtc tgtctacatt gaattggctg cttgtgaatt ttctgagacg gatgtaacaa
 +
      781 atactgaaca tcatcaaccc agtaataatg atttgaacac cactgagaag cgtgcagctg
 +
      841 agaggcatcc agaaaagtat cagggtgaag cagcatctgg gtgtgagagt gaaacaagcg
 +
      901 tctctgaaga ctgctcaggg ctatcctctc agagtgacat tttaaccact cagcagaggg
 +
      961 ataccatgca acataacctg ataaagctcc agcaggaaat ggctgaacta gaagctgtgt
 +
    1021 tagaacagca tgggagccag ccttctaaca gctacccttc catcataagt gactcttctg
 +
    1081 cccttgagga cctgcgaaat ccagaacaaa gcacatcaga aaaagcagta ttaacttcac
 +
    1141 agaaaagtag tgaataccct ataagccaga atccagaagg cctttctgct gacaagtttg
 +
    1201 aggtgtctgc agatagttct accagtaaaa ataaagaacc aggagtggaa aggtcatccc
 +
    1261 cttctaaatg cccatcatta gatgataggt ggtacatgca cagttgctct gggagtcttc
 +
    1321 agaatagaaa ctacccatct caagaggagc tcattaaggt tgttgatgtg gaggagcaac
 +
    1381 agctggaaga gtctgggcca cacgatttga cggaaacatc ttacttgcca aggcaagatc
 +
    1441 tagagggaac cccttacctg gaatctggaa tcagcctctt ctctgatgac cctgaatctg
 +
    1501 atccttctga agacagagcc ccagagtcag ctcgtgttgg caacatacca tcttcaacct
 +
    1561 ctgcattgaa agttccccaa ttgaaagttg cagaatctgc ccagagtcca gctgctgctc
 +
    1621 atactactga tactgctggg tataatgcaa tggaagaaag tgtgagcagg gagaagccag
 +
    1681 aattgacagc ttcaacagaa agggtcaaca aaagaatgtc catggtggtg tctggcctga
 +
    1741 ccccagaaga atttatgctc gtgtacaagt ttgccagaaa acaccacatc actttaacta
 +
    1801 atctaattac tgaagagact actcatgttg ttatgaaaac agatgctgag tttgtgtgtg
 +
    1861 aacggacact gaaatatttt ctaggaattg cgggaggaaa atgggtagtt agctatttct
 +
    1921 gggtgaccca gtctattaaa gaaagaaaaa tgctgaatga gcatgatttt gaagtcagag
 +
    1981 gagatgtggt caatggaaga aaccaccaag gtccaaagcg agcaagagaa tcccaggaca
 +
    2041 gaaagatctt cagggggcta gaaatctgtt gctatgggcc cttcaccaac atgcccacag
 +
    2101 atcaactgga atggatggta cagctgtgtg gtgcttctgt ggtgaaggag ctttcatcat
 +
    2161 tcacccttgg cacaggtgtc cacccaattg tggttgtgca gccagatgcc tggacagagg
 +
    2221 acaatggctt ccatgcaatt gggcagatgt gtgaggcacc tgtggtgacc cgagagtggg
 +
    2281 tgttggacag tgtagcactc taccagtgcc aggagctgga cacctacctg ataccccaga
 +
    2341 tcccccacag ccactactga

Current revision

How to be a Breast Cancer Detective.

Lesson Overview

Briefly describe the lesson here.

The lesson begins with interpreting familial pedigree charts and continues with steps in the identification of the genes responsible for some breast cancers. It continues with searching for the BRCA1 gene and analysis and identification of the coding sequences through bioinformatics techniques.

Goals and Objectives

1) understand the process by which genes of parents are transferred to their offspring.

2) understand the difference between a dominant and recessive trait and understand how the presence of a trait may effect the physical characteristics of an individual.

3) read and/or construct a pedigree chart mapping a specific trait in a family.

4) determine the probability of a certain phenotype being expressed in a pedigree chart

5) Understand the uses of Biotechnology and Bioinformatics in understanding cancer

6) Learn to retrieve information from a Bioinformatics source.

Common Misconceptions


Students may not understand the importance and relevance of molecular genetics to research on today's world. Many students are not aware of the world of Bioinformatics.


The Lesson

Preparation Before class

Part 1. PowerPoint to accompany lesson, Introduction to Pedigree charts (handout),Highlighters, pens, pencils, books, chalkboard, and 1 computer with LCD projector and sound system

Part 2: Computer classroom; one computer per student, LCD projector,sound system

During class

Time required

Part 1- 45 minutes Part 1: Predicting the Breast cancer gene through Pedigree charts 

       Practice drawing a pedigree chart

Begin by using PowerPoint presentation which contains links to other resources listed below:

Mary Claire King: Finding brca1 and 2 by pedigree- http://www.dnai.org/media/a/king296¬04.swf

Mark Skolnick: Finding the breast cancer gene brca1-http://www.dnai.org/media/a/skolnick298_06.swf

Mary Claire king: Can women be tested for breast cancer? http://www.dnaiorg/media/a/king295_08.swf


Time required

Part 2 -2 45 minute periods

How do mutations form? View film clip http://www.insidecancer.org/?s=A,1,7

1. Reading and Discussion of Biotechnology and Bioinformatics based on the reading below: 

      Biotechnology’s Relation to Cancer Research

Biotechnology has applied many of the techniques of cell biology, molecular biology, and protein chemistry to cancer research and treatment over the years. These developments include:the ability to create vaccines; the identification of protein markers for detecting and diagnosing cancers; the purification of interleukin and other immune-system enhancers; the preparation of bone marrow cells for transplants;the manipulation of natural products like taxol (from the Pacific yew tree) to make improved compounds that interfere with cell division. New diagnostic abilities and drugs will result from related advances in genomics, proteomics, computer modeling, and bioinformatics. Genomics (the study of the genes on the chromosomes) is showing the exact chromosomal defect involved in cancers and pinpointing how key genes malfunction. Proteomics (the study of the proteins made by the genes) will define the function of the protein made by the defective genes in the cascade of cancer events. Computer modeling allows proteins to be considered in a variety of new lights, including indentifying potential docking clefts for the introduction of small molecules or proteins that might interfere with cancer causing protein functions. Bioinformatics and combinatorial chemistry allow scientists to sort through molecules (both natural and synthetic) that interfere with cancer’s progress in a variety of ways. In addition, the ability to measure many different constituents (such as the DNA, RNA, and proteins) in both normal cells and cancer cells, will enable a more systemic genetic classification of cancer. Bioinformatics can discern patterns in those measurements to help better understand how cancer cells behave and how they react to different treatments. (from Biotechnology and You,vol.11,issue 1)

2. Finding the Brca 1 gene using bioinformatics


Student Handouts for the Lesson Plan

Instruction sheet on steps in finding genes and using the NCBI website

Alternative Assessments

Poster of Breast Cancer Pedigree with correct key

Bioinformatics project using tools and websites found in this lesson.


Suggestions for Extended Learning

• Webster K Cavenee and Raymond L. White, “The Genetic Basis of Cancer,” Scientific American, March 1995.

• Matt Ridley, Chapter 17 in Genome: Autobiography of a Species in 23 Chapters

• Robert Weinberg, One Renegade Cell

• Time and Newsweek cover articles on cancer topics

• American Cancer Society: http://www.cancer.org

• National Cancer Institute: http://www.nci.nih.gov

• National Childhood Cancer Foundation: http://www.nccf.org

• Oncolink: http://www.oncolink.upenn.edu

• NOVA’s program on Judah Folkman, Cancer Warrior: http://pbs.org/wgbh/ nova/cancer


Glossary

Allele—an alternate form of a gene.

Chromosome—structure in the cell nucleus that stores and transmits genetic information.

Gene—unit of heredity.

Genotype—Allelic status of an organism for a genetic trait.

Heterozygous—having different alleles of a gene.

Homozygous—having indistinguishable alleles of a gene.

Pedigree—diagram showing the expression of a specific characteristic and the biological relationships among members of a family, often of several generations.

Phenotype—the observable organism, the expression of a genetic trait.

Bioinformatics- the science of using databases to enter or research genetic information on the WWW.

NCBI-National Center for Biotechnology Information

Biotechnology-the scientific manipulation of living organisms, especially at the molecular genetic level, to produce useful products.

CDS-Coding sequence


Education Standards 

• Living systems also have different levels of organization--for example, cells, tissues, organs, organisms, populations, and communities. Within these systems, interactions between components occur. Further, systems at different levels of organization can manifest different properties and functions.

• Models are tentative schemes or structures that correspond to real objects, events, or classes of events, and that have explanatory power. Models help scientists and engineers understand how things work. Models take many forms, including physical objects, plans, mental constructs, mathematical equations, and computer simulations.

• Cells have particular structures that underlie their functions. Every cell is surrounded by a membrane that separates it from the outside world. Inside the cell is a concentrated mixture of thousands of different molecules which form a variety of specialized structures that carry out such cell functions as energy production, transport of molecules, waste disposal, synthesis of new molecules, and the storage of genetic material.

• Cells store and use information to guide their functions. The genetic information stored in DNA is used to direct the synthesis of the thousands of proteins that each cell requires.

• In all organisms, the instructions for specifying the characteristics of the organism are carried in DNA, a large polymer formed from subunits of four kinds (A, G, C, and T). The chemical and structural properties of DNA explain how the genetic information that underlies heredity is both encoded in genes (as a string of molecular "letters") and replicated (by a templating mechanism). Each DNA molecule in a cell forms a single chromosome.

• Recognize and analyze alternative explanations and models, reviewing current scientific understanding.

Teacher Answer Key

Part 2: Bioinformatics Brca 1 gene:

Questions

1.What is a locus? A position in the genome where a particular gene is found

2.How many “bp” base pairs are found in this locus? 3759 base pairs

3.Where is the gene found? mRNA Homo Sapiens

4.What does the gene encode for? 

           This gene encodes a nuclear phosphoprotein that plays a
           role in maintaining genomic stability and acts as a tumor
           suppressor. The encoded protein combines with other tumor
           suppressors, DNA damage sensors, and signal transducers to form a
           large multi-subunit protein complex known as BASC for
           BRCA1-associated genome surveillance complex.

CDS of Human Breast cancer gene 

                                tggatttat ctgctcttcg cgttgaagaa gtacaaaatg
     241 tcattaatgc tatgcagaaa atcttagagt gtcccatctg tctggagttg atcaaggaac
     301 ctgtctccac aaagtgtgac cacatatttt gcaaattttg catgctgaaa cttctcaacc
     361 agaagaaagg gccttcacag tgtcctttat gtaagaatga tataaccaaa aggagcctac
     421 aagaaagtac gagatttagt caacttgttg aagagctatt gaaaatcatt tgtgcttttc
     481 agcttgacac aggtttggag tatgcaaaca gctataattt tgcaaaaaag gaaaataact
     541 ctcctgaaca tctaaaagat gaagtttcta tcatccaaag tatgggctac agaaaccgtg
     601 ccaaaagact tctacagagt gaacccgaaa atccttcctt gcaggaaacc agtctcagtg
     661 tccaactctc taaccttgga actgtgagaa ctctgaggac aaagcagcgg atacaacctc
     721 aaaagacgtc tgtctacatt gaattggctg cttgtgaatt ttctgagacg gatgtaacaa
     781 atactgaaca tcatcaaccc agtaataatg atttgaacac cactgagaag cgtgcagctg
     841 agaggcatcc agaaaagtat cagggtgaag cagcatctgg gtgtgagagt gaaacaagcg
     901 tctctgaaga ctgctcaggg ctatcctctc agagtgacat tttaaccact cagcagaggg
     961 ataccatgca acataacctg ataaagctcc agcaggaaat ggctgaacta gaagctgtgt
    1021 tagaacagca tgggagccag ccttctaaca gctacccttc catcataagt gactcttctg
    1081 cccttgagga cctgcgaaat ccagaacaaa gcacatcaga aaaagcagta ttaacttcac
    1141 agaaaagtag tgaataccct ataagccaga atccagaagg cctttctgct gacaagtttg
    1201 aggtgtctgc agatagttct accagtaaaa ataaagaacc aggagtggaa aggtcatccc
    1261 cttctaaatg cccatcatta gatgataggt ggtacatgca cagttgctct gggagtcttc
    1321 agaatagaaa ctacccatct caagaggagc tcattaaggt tgttgatgtg gaggagcaac
    1381 agctggaaga gtctgggcca cacgatttga cggaaacatc ttacttgcca aggcaagatc
    1441 tagagggaac cccttacctg gaatctggaa tcagcctctt ctctgatgac cctgaatctg
    1501 atccttctga agacagagcc ccagagtcag ctcgtgttgg caacatacca tcttcaacct
    1561 ctgcattgaa agttccccaa ttgaaagttg cagaatctgc ccagagtcca gctgctgctc
    1621 atactactga tactgctggg tataatgcaa tggaagaaag tgtgagcagg gagaagccag
    1681 aattgacagc ttcaacagaa agggtcaaca aaagaatgtc catggtggtg tctggcctga
    1741 ccccagaaga atttatgctc gtgtacaagt ttgccagaaa acaccacatc actttaacta
    1801 atctaattac tgaagagact actcatgttg ttatgaaaac agatgctgag tttgtgtgtg
    1861 aacggacact gaaatatttt ctaggaattg cgggaggaaa atgggtagtt agctatttct
    1921 gggtgaccca gtctattaaa gaaagaaaaa tgctgaatga gcatgatttt gaagtcagag
    1981 gagatgtggt caatggaaga aaccaccaag gtccaaagcg agcaagagaa tcccaggaca
    2041 gaaagatctt cagggggcta gaaatctgtt gctatgggcc cttcaccaac atgcccacag
    2101 atcaactgga atggatggta cagctgtgtg gtgcttctgt ggtgaaggag ctttcatcat
    2161 tcacccttgg cacaggtgtc cacccaattg tggttgtgca gccagatgcc tggacagagg
    2221 acaatggctt ccatgcaatt gggcagatgt gtgaggcacc tgtggtgacc cgagagtggg
    2281 tgttggacag tgtagcactc taccagtgcc aggagctgga cacctacctg ataccccaga
    2341 tcccccacag ccactactga
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